A developmental switch of gene expression in the barley seed …€¦ · 26/02/2016 · The deduced HvVP1 protein sequence from cv Igri is more than 99% 161 identical with those

1

A developmental switch of gene expression in the barley seed mediated 1 by HvVP1 (Viviparous-1) and HvGAMYB interactions 2

3

Zamira Abraham*, Raquel Iglesias-Fernández*, Manuel Martínez, Isabel Díaz, 4 Pilar Carbonero, Jesús Vicente-Carbajosa+ 5

*Equal authors 6

Centro de Biotecnología y Genómica de Plantas (UPM-INIA), and E.T.S.I. 7 Agrónomos, Campus de Montegancedo, Universidad Politécnica de Madrid, 8 Pozuelo de Alarcón, 28223-Madrid, Spain 9 10

Short Title: HvVP1 effects on HvGAMYB in seed development 11

12

Keywords: Barley Viviparous-1 (HvVP1), HvGAMYB, BPBF (HvDOF24), Hor2 13 gene, α-amylase gene, seed gene regulation, maturation, germination, 14 transcriptional activation, transcriptional repression 15

1 This work was financially supported by MINECO, Spain (Project BFU2013-16

49665-EXP; p.i. J. V-C) and by MICINN (Project BFU2009-11809; p.i. P.C). 17

+Address correspondence to [email protected] 18

The author responsible for distribution of materials integral to the findings 19 presented in this article in accordance with the policy, described in the 20 instructions for authors (http://www.plantphysiolo.org), is Jesús Vicente-21 Carbajosa ([email protected]). 22

Z.A, R.I-F, M.M and I.D performed the experiments and P.C and J.V-C 23 conceived and supervised the experiments; J.V-C, P.C and R.I-F wrote the 24 article with the contributions from other authors. 25

26

Email addresses: 27

Z.A.: [email protected] 28

R.I.F.: [email protected] 29

M.M.: [email protected] 30

I.D.: [email protected] 31

P.C.: [email protected] 32

Plant Physiology Preview. Published on February 26, 2016, as DOI:10.1104/pp.16.00092

Copyright 2016 by the American Society of Plant Biologists

https://plantphysiol.orgDownloaded on December 25, 2020. - Published by Copyright (c) 2020 American Society of Plant Biologists. All rights reserved.







https://plantphysiol.org







2

SUMMARY 33

Accumulation of storage compounds in the starchy endosperm of developing 34

cereal seeds is highly regulated at the transcriptional level. These compounds, 35

mainly starch and proteins, are hydrolyzed upon germination to allow seedling 36

growth. The transcription factor HvGAMYB is a master activator both in the 37

maturation phase of seed development and upon germination, acting in 38

combination with other transcription factors. However, the precise mechanism 39

controlling the switch from maturation to germination programs remains unclear. 40

We report here the identification and molecular characterization of barley 41

HvVP1 (Viviparous-1), orthologous to ABI3 from Arabidopsis. HvVP1 transcripts 42

accumulate in the endosperm and the embryo of developing seeds at early 43

stages, and in the embryo and aleurone of germinating seeds up to 24 hours of 44

imbibition (hoi). In transient expression assays, HvVP1 controls the activation of 45

Hor2 and Amy6.4 promoters exerted by HvGAMYB. HvVP1 interacts with 46

HvGAMYB in yeast and in the plant nuclei, hindering its interaction with other 47

transcription factors involved in seed gene expression programs, like BPBF. 48

Similarly, this interaction leads to a decrease in the DNA-binding of HvGAMYB 49

and BPBF to their targets sequences. Our results indicate that the HvVP1 50

expression pattern controls the full Hor2 expression activated by GAMYB and 51

BPBF in the developing endosperm, and the Amy6.4 activation in post-52

germinative reserve mobilization mediated by GAMYB. All these data 53

demonstrate the participation of HvVP1 in antagonistic gene expression 54

programs and support its central role as a gene expression switch during seed 55

maturation and germination. 56



3

INTRODUCTION 57

Seed development after fertilization has been classically divided into two 58

major phases: zygotic embryogenesis and maturation (Goldberg et al., 1994). 59

The maturation phase (MAT) is characterized by gene expression programs 60

devoted to the synthesis and accumulation of reserve compounds, among them 61

Seed Storage Proteins (SSPs) and, later on, to the acquisition of desiccation 62

tolerance, with an important role for the Late Embryogenic Abundant proteins 63

(LEAs; Vicente-Carbajosa and Carbonero, 2005). In most species of the 64

Spermatophyta, seeds acquire a quiescent state at the end of the maturation 65

phase that is resumed by the germination process (GER), which is generally 66

accepted to comprise the germination sensu stricto from seed imbibition to root 67

emergence that is followed by reserve mobilization. These two distinct phases 68

are independently regulated and influenced by genetic and environmental 69

factors (Bewley, 1997; Finch-Savage and Leubner-Metzger, 2006; Nonogaki et 70

al., 2007; Holdsworth et al., 2008; Iglesias-Fernández et al., 2011a; González-71

Calle et al., 2015). During early post-germination (reserve mobilization), several 72

genes encoding hydrolytic enzymes (i.e.: α-amylases, proteases and lipases) 73

catalyze the hydrolysis of carbohydrates, proteins and lipids present in the seed 74

storage tissues, such as the endosperm of Monocotyledonous species and the 75

cotyledons of Dicotyledonous ones, with the aim of nourishing the growing 76

embryo before attaining its full photosynthetic capacity (Pritchard et al., 2002; 77

González-Calle et al., 2014; Iglesias-Fernández et al., 2014). 78

The comprehensive molecular and genetic mechanisms controlling seed 79

development and germination still remain elusive, due mainly to the many 80

environmental and intrinsic parameters influencing it. From a physiological 81



4

point of view, it is generally accepted that an increased Abscisic 82

Acid/Gibberellins (ABA/GA) ratio determines the progression of seed 83

maturation, by promoting the expression of SSPs, LEAs, Heat Shock Proteins 84

(HSPs), and Anthocyanin biosynthesis genes (Reidt et al., 2000; Koornneef et 85

al., 2002; Vicente-Carbajosa and Carbonero, 2005; Braybrook and Harada, 86

2008; Finkelstein et al., 2008). By contrast, an increased GA/ABA ratio is the 87

most important factor for the integration of the environmental and intrinsic 88

signals for the occurrence of germination (Kucera et al., 2005). This enhanced 89

GA/ABA balance raises the expression of hydrolase genes such as those 90

encoding Cell Wall Remodeling Enzymes (CWREs), involved in the weakening 91

of the embryo surrounding tissues during germination sensu stricto, and of 92

Reserve Mobilization Enzymes in post-germination (RMEs; Gubler et al., 1995; 93

1999; Mena et al., 2002; Iglesias-Fernández et al., 2011b, c; González-Calle et 94

al., 2015). 95

In cereal seeds, the transcription factor (TF) of the B3 family Viviparous-1 96

(VP1), orthologous to ABA-Insensitive-3 (ABI3) from Arabidopsis thaliana, is a 97

key component of the ABA signaling and is required for expression of the 98

maturation program (McCarty et al., 1991; Parcy et al., 1994; Jones et al., 1997; 99

Nambara et al. 2000; Suzuki et al.,2001; Lara et al., 2003; Zeng et al., 2003; To 100

et al., 2006; Swaminathan et al., 2008; Yan et al., 2014). The Viviparous-1 101

mutants, like vp1 in Zea mays or abi3 in Arabidopsis thaliana, are characterized 102

by the incapacity of entering the quiescent state at the end of the maturation 103

phase, and by displaying premature features of the subsequent germination and 104

post-germination phases, leading to a precocious germination of the seeds still 105

in the mother plant (pre-harvest sprouting; McCarty et al., 1989; Gubler et al., 106



5

2005). It has been previously described that the VP1 TF, through its binding to 107

the specific RY cis-element (5´-CATGCA-3´) and through protein-protein 108

interactions with other TFs of the bZIP family, regulates the expression of seed 109

maturation specific genes, such as those encoding the Em protein in the wheat 110

caryopsis and the C1 (MYB-like) transcription factor, a key regulator of the 111

anthocyanin biosynthesis pathway in maize (Paz-Ares et al., 1986; Hattori et al., 112

1992; Vasil et al., 1995). VP1 has been also associated with repression of post-113

germination genes, since in the maize vp1 mutant, alpha-amylase activity is 114

precociously observed in the kernels while still in the corn cob, indicating that all 115

necessary factors for its activation are already present during the maturation 116

phase, and in support that the VP1 protein must be preventing this hydrolytic 117

activity (Hoecker et al., 1999; Rodríguez et al., 2015). 118

In the last twenty years, an important number of barley TFs, such as GAMYB, 119

BPBF (HvDOF24), SAD (HvDOF23) and BLZ2 (a bZIP orthologous to the maize 120

OPAQUE-2), have been characterized as central regulators of gene expression 121

in the seed maturation and post-germination (reserve mobilization) programs 122

(Gubler et al, 1995; Mena et al., 1998; 2002; Oñate et al., 1999; Díaz et al., 123

2002; Isabel-LaMoneda et al., 2003). However, their possible relationship with 124

the barley VP1 has not been investigated, although similar interactions have 125

been reported in other cereal species (Hill et al., 1996; Hobo et al., 1999), and 126

for ABI3 and AtbZIP10/AtbZIP25 (BLZ2 orthologs) with a role in activating the 127

expression of SSP genes upon seed maturation in A. thaliana (Lara et al., 128

2003). 129

130



6

In this work, the barley VP1 gene (HvVP1) has been characterized and 131

its expression localized, by mRNA in situ hybridization experiments, to the 132

embryo and the endosperm of barley seeds, both during maturation and upon 133

germination. In transient expression experiments in immature barley 134

endosperms and in aleurone layers of germinating seeds, HvVP1 interferes with 135

the transcriptional activation exerted by GAMYB on the promoters of the genes 136

Hor2 and Amy6.4, encoding a B-hordein (SSP) and a high-pI α-amylase, 137

respectively. By yeast 2-hybrid (Y2H) and Bimolecular Fluorescence 138

Complementation (BiFC) assays, the HvVP1-GAMYB protein interaction has 139

been confirmed in vivo and in plant nuclei. Interestingly, the presence of HvVP1 140

not only diminishes the GAMYB-BPBF protein interaction in yeast three hybrid 141

(Y3H) assays, but HvVP1 also decreases the binding affinities of GAMYB and 142

BPBF for their corresponding cis-elements in the promoters of the Hor2 and 143

Amy6.4 genes in Electrophoretic Mobility Shift Assays (EMSAs). All these data 144

indicate a central role for HvVP1 as a gene expression switch at key stages of 145

seed maturation and germination. 146

147

RESULTS 148

The HvVP1 Sequence Identification and Phylogenetic Dendrogram 149

The sequence and structure of the predicted VIVIPAROUS-1 gene 150

(HvVP1), isolated from a barley (cv Igri) genomic library (Supplemental Fig. S1) 151

was confirmed by sequencing the corresponding Open Reading Frame (ORF) 152

derived from a PCR amplified cDNA from developing barley seeds (20 days 153

after pollination, dap). This sequence was compared with other VP1 sequences 154

from different barley cvs (Igri, Morex, Haruna Nijo) and with its orthologs in 155



7

other Gramineae deposited in public databases: Triticum aestivum (TaVP1), 156

Triticum turgidum (TtVP1), Triticum monococum (TmVP1), Brachypodium 157

distachyon (BdVP1) and Oryza sativa (OsVP1), and with AtABI3 from 158

Arabidopsis thaliana. The HvVP1 gene is a single copy gene (Supplemental 159

Fig. S2). The deduced HvVP1 protein sequence from cv Igri is more than 99% 160

identical with those from cv. Morex (International Barley Genome Sequencing 161

Consortium, 2012), and from cv Haruna Nijo (Matsumoto et al., 2011), and with 162

the VP1 partial sequence of cv. Himalaya in the Genbank (AAO06117.1; 163

Casaretto and Ho, 2003). A phylogenetic tree including these sequences has 164

been constructed (Fig. 1A) and HvVP1 is included in the same branch as those 165

of the Triticeae tribe species with a bootstrap value of 100%. This phylogenetic 166

tree is further supported by the occurrence of common motives (MEME 167

analysis, Fig. 1B; Table 1). All sequences share motives 1, 2, 3, 4, 5, 14 and 15, 168

with the exception of motif 3 that is only partially conserved in AtABI3. 169

According to Nakamura and Toyama (2001), motif 3 corresponds to the 170

activation domain (A), motives 15 and 2 form the B1 domain, motif 5 matches 171

with the B2 domain, and motives 1, 4 and 14 integrate the B3 (DNA-binding) 172

domain. VP1 proteins from members of the Triticeae tribe (TaVP1, TtVP1, 173

TmVP1 and HvVP1) share motives 8, 9, 10, 12, 16, 19 and 20, and all the VP1 174

proteins in the Poaceae genomes compared, share motives 6, 13, 17, 18 and 175

21. 176

The deduced amino-acid sequences encoded by these VP1 genes have 177

nuclear localization signals (RKKR), molecular weights of 72-75 kDa and 178

isoelectric points between 6.4-8.8 (Supplemental Table 1). The HvVP1 gene 179

contains six exons (solid bars) and five introns (lines; Fig. 2A) as determined by 180



8

comparing the genomic and cDNA clones. While domains A, B1 and B2 181

(containing the nuclear localization signal RKKR; Graeber et al., 2010) are 182

encoded in the first exon, the B3 DNA-binding domain is translated from exons 183

II-VI. 184

185

Activation properties of the HvVP1 protein 186

To investigate the activation capacity of HvVP1 in yeast, a series of 187

constructs containing different regions of the HvVP1 ORF was prepared (Fig. 188

2B). These regions were cloned as translational fusions to the GAL4 DNA 189

binding domain into a yeast expression plasmid and tested as effectors for their 190

capacity to transactivate two alternative reporter genes: LacZ encoding a β-191

galactosidase assayed in liquid cultured cells, and HIS3 that promotes yeast 192

growth in a histidine depleted agar medium. Both, in qualitative (His-) and in 193

quantitative (β-Gal, Miller Units, M.U.) assays, HvVP1 acts as a potent activator 194

in yeast (~450 β-Gal M.U.). However, when domain A is deleted, the reporter 195

expression disappears, indicating that this region is essential for HvVP1 being a 196

transcriptional activator in yeast. 197

198

HvVP1 is expressed in the Embryo and in the Endosperm during Seed 199

Maturation and Germination 200

Cereal VP1 proteins, as Arabidopsis ABI3, belong to a sub-family of B3 201

TFs controlling seed gene expression (Swaminathan et al., 2008). 202

Consequently, it is expected that the HvVP1-mRNA should be expressed during 203



9

barley seed maturation and germination. To test this, total RNA was isolated 204

from developing de-embrionated seeds (endosperms at 5, 10, 15, 20 dap), 205

immature (20 dap) and mature dry embryos, and from aleurones and embryos 206

of germinating seeds at different times of imbibition (8, 16, 24, 48, 72 hours of 207

imbibition, hoi). Northern blot analysis (Fig. 3) shows that the HvVP1 transcripts 208

are found both in developing endosperms, attaining maximum levels at 5 dap, 209

decreasing thereafter (20 dap), and in immature and mature embryos, being 210

more abundant at the immature stage (Fig. 3A). The HvVP1 transcripts are 211

expressed upon seed germination in the aleurone layer, reaching a peak at 24 212

hoi, and also in embryos (Fig. 3B). mRNA in situ hybridization experiments 213

during seed maturation (20 dap) and upon germination (24 hoi) reveal that 214

HvVP1 transcripts are present throughout the seed (embryo and endosperm) 215

during maturation, while its expression is restricted to the embryo and the 216

aleurone layer upon germination (Figs. 3A, 3B). The HvVP1 transcripts are not 217

found in leaves and roots of barley adult plants (our data not shown). 218

219

HvVP1 counteracts the HvGAMYB Trans-Activation exerted on 220

Endosperm specific Genes during Seed Maturation and Germination 221

The HvGAMYB is a barley MYB-R2R3 protein that is a transcriptional 222

activator of seed storage protein genes (B-hordeins: HvHor2) during seed 223

maturation and of hydrolase genes (α-amylases: HvAmy6.4) needed for reserve 224

mobilization in post-germination (Gubler et al., 1995; 1999; Díaz et al., 2002). 225

Taking into account that both HvVP1 (Fig. 3) and HvGAMYB share 226

similar spatial expression patterns in the seed, as occurs with other important 227



10

seed specific TFs (Supplemental Fig. S4), transient expression assays were 228

done by co-bombardment in barley developing endosperms and in germinating 229

aleurones to investigate their mutual effects. In Figure 4A, the schematic 230

description of the constructs used in the trans-activation study is shown. The 231

transcription factors HvVP1 and HvGAMYB were used as effectors. As reporter 232

constructs, the HvHor2 promoter (-560 bp upstream of the ATG initiation codon) 233

and the HvAmy6.4 gene promoter (-174 bp) were fused to the GUS reporter 234

gene (PHor2:uidA and PAmy6.4:uidA). As shown in Fig. 4B, while co-bombardment 235

of developing barley endosperms with HvVP1 and PHor2:uidA reduces GUS 236

activity to half that driven by the PHor2:uidA construct alone, co-transfection with 237

HvGAMYB and PHor2:uidA provokes an increment of GUS activity of ~5-fold. 238

When both TFs (HvVP1 and HvGAMYB) are co-bombarded together with 239

PHor2:uidA, the positive trans-activation of PHor2:uidA exerted by HvGAMYB 240

diminishes drastically (Fig. 4B). Similarly, Fig. 4C shows that while co-241

bombardment of HvVP1 with PAmy6.4:uidA diminishes GUS activity to half that 242

driven by the reporter gene alone (PAmy6.4:uidA), co-transfection of HvGAMYB 243

and PAmy6.4:uidA increases GUS activity more than ~8-fold, and co-244

bombardment of both TFs reduces the GUS activity of the reporter construct to 245

half that obtained when HvGAMYB is the only transfected TF. This repressor 246

effect of HvVP1 is independent of the presence of GA (1μM) added to the 247

incubation medium (Supplemental Fig. S5). In summary, HvVP1 is a 248

transcriptional repressor of maturation and germination seed genes. 249

250

HvVP1 interacts with HvGAMYB in the Yeast Two-Hybrid System and in 251

Plant Nuclei 252



11

To explore whether the repression exerted by the HvVP1 TF on the 253

trans-activation mediated by HvGAMYB on seed specific genes during 254

maturation and germination could be due to a direct HvVP1-HvGAMYB protein-255

protein interaction, yeast two-hybrid and bimolecular fluorescence 256

complementation assays have been done. 257

LacZ and HIS3 reporter activities were measured in yeast two-hybrid 258

assays (Fig. 5). The HvVP1-N terminal cDNA, encoding 464 amino-acid 259

residues, spanning domains A, B1 and B2 (Fig. 2A), was translationally fused to 260

the yeast GAL4 activation domain (AD), and the full-length cDNA of HvGAMYB 261

was fused to the GAL4 binding domain (BD; Fig. 5B). In Fig. 5C, yeast cells 262

(strain SFY526) transformed with the GAL4BD-HvGAMYB construct show 263

detectable background levels of LacZ reporter activity, and this activity sharply 264

increases (~400 β-Gal M.U.) when this strain is co-transformed with the 265

GAL4AD-HvVP1-N construct. Expression of the HIS3 gene (strain HF7C), 266

conferring histidine auxotrophy, was used to confirm the interaction between 267

HvGAMYB and HvVP1N (Fig. 5C). Yeast cells containing GAL4BD-HvGAMYB 268

and the GAL4AD-Ø constructs are not able to grow in 3-aminotriazol (3-AT) 269

concentrations higher than 30 mM. However, when transformed with GAL4BD-270

HvGAMYB and GAL4AD-HvVP1N constructs, yeast cells grow even at 60 mM 271

of 3-AT. All these results support that the HvGAMYB and HvVP1 proteins 272

interact in vivo. 273

In order to corroborate the HvVP1-HvGAMYB interaction, bimolecular 274

fluorescence complementation experiments were done in planta. With this aim, 275

HvVP1 and HvGAMYB ORFs were translationally fused to the N- and C- 276

terminal fragments, respectively, of the green fluorescent protein encoding gene 277



12

(GFP; Díaz et al., 2005) and used for co-bombardment experiments in onion 278

epidermal cells. Microscopic observations show that the GFP fluorescence is 279

reconstituted and targeted to the nucleus, indicating that HvVP1 and HvGAMYB 280

proteins interact in plant nuclei (Fig. 6). As expected, there is no detectable 281

GFP when the GFP fragments are bombarded alone (data not shown). 282

283

HvVP1 interferes with the HvGAMYB-BPBF Interaction in vivo and with 284

their DNA Binding in Electrophoretic Mobility Shift Assays (EMSA) 285

Since we showed that HvVP1 and GAMYB proteins can physically 286

interact, we wanted to determine whether the HvVP1 protein could interfere with 287

the binary complex formed by HvGAMYB and BPBF-HvDOF24, previously 288

reported (Díaz et al., 2002) For this purpose a yeast three-hybrid (Y3H) system 289

was used, based on the pBridgeTM vector, which allows the simultaneous 290

expression of two proteins. In this case, the Gal4BD-BPBF translational fusion 291

and the HvVP1-ORF, the latter conditionally expressed under the control of the 292

Met25 promoter (PMet25) that responds to the methionine (Met) supply in the 293

medium. The Gal4AD-HvGAMYB construction was used as the third component 294

of the system (Fig. 7A). In this Y3H system and in the absence of Met in the 295

yeast growth medium, the joint expression of HvGAMYB-AD and BPBF-BD 296

increases the reporter gene activity (>80 β-Gal M.U.), compared to the 297

expression of BPBF alone (~40 β-Gal M.U.), as expected for a positive 298

interaction between the two proteins. However, this activity diminishes when 299

HvVP1 is present (~60 β-Gal M.U.; Fig. 7B) suggesting that HvVP1 interferes 300

with the interaction between HvGAMYB and BPBF. 301



13

The specific DNA-protein interaction between BPBF (HvDOF24) and its 302

corresponding cis motif (Prolamine Box, PB: 5´-TGTAAAG-3´), and between 303

HvGAMYB and the MYB-R2R3 box (5´-TAACAAC-3´) at the promoter region of 304

the HvHor2 gene have been previously reported (Mena et al., 1998; Díaz et al., 305

2002). The potential effect of HvVP1 in modifying these protein-DNA 306

interactions was investigated by Electrophoretic Mobility Shift Assays (EMSA; 307

Fig. 7C). HvVP1, HvGAMYB and BPBF proteins produced in E. coli were 308

assayed for their binding activities using two [32P]-labeled DNA probes 309

containing the MYB-R2R3 and the Prolamine Box binding sites, derived from 310

the HvHor2 promoter (Fig. 7C). While a clear retarded band is observed when 311

HvGAMYB or BPBF) protein extracts are incubated with their respective probes, 312

the addition of increasing amounts of HvVP1 protein to the reactions leads to 313

fainter, or even to the absence of retarded bands. Notably, HvVP1 does not 314

bind to any of the probes used containing the PB/GLM motives (target binding 315

sites of BPBF and BLZ2, respectively) or the MYB-R2R3 box (target binding site 316

of GAMYB) in support of its direct interference with the binding of these 317

proteins. Interestingly, modification of DNA-binding by HvVP1 seems to be 318

specific, since the intensity of the retarded bands in EMSAs for SAD 319

(HvDOF23) was decreased in the presence of HvVP1, while the contrary effect 320

(enhancement) was observed for the bZIP protein BLZ2 (Supplemental Fig. S6) 321

322

HvVP1 counteracts the Transcriptional Activation of BPBF on the HvHor2 323

gene promoter 324

BPBF is a barley DOF transcription factor that acts as a transcriptional 325

activator of the endosperm specific gene HvHor2 during seed maturation (Mena 326



14

et al., 1998), and the BPBF transcripts are abundant during the seed maturation 327

and germination phases (Supplemental Fig. S4). 328

As BPBF shares similar spatial expression patterns during seed 329

development with HvVP1, and the binding of its encoded protein for the PB box 330

is drastically decreased by HvVP1 (EMSA in Fig. 7C), we decided to perform 331

transient expression assays by particle bombardment in developing barley 332

endosperms to explore the in planta effects of the coexpression of both TFs. 333

Figure 8A schematically shows the constructs used in the transactivation study, 334

where BPBF and HvVP1 were tested as effectors on GUS reporter activity 335

driven by the PHor2:uidA construct. As shown in Fig. 8B, co-bombardment of 336

barley developing endosperms with the HvVP1 effector and PHor2:uidA reporter, 337

diminishes GUS activity to half that of PHor2:uidA alone, whereas co-transfection 338

with BPBF and PHor2:uidA provokes an increment of GUS activity of ~7-fold. 339

When both HvVP1 and BPBF are co-bombarded, the positive PHor2:uidA trans-340

activation exerted by BPBF is reduced to ~ half. This result demonstrates that 341

HvVP1 counteracts the activation of BPBF upon the HvHor2 gene in developing 342

barley endosperms. 343

344

DISCUSSION 345

In this work, we have identified the Viviparous-1 (HvVP1) gene of 346

Hordeum vulgare cv. Igri and explored its function in the control of gene 347

expression programs during seed development and germination. In particular, 348

we address the role of HvVP1 through its interaction with HvGAMYB, a key 349

transcription factor that participates in the regulation of both MAT and GER 350



15

genes in the barley seed. Our work unveils important features of HvVP1 action 351

derived from its precise spatio-temporal expression patterns in the seed, and its 352

capacity to modulate regulatory complexes in which HvGAMYB participates. We 353

propose a general model (Figure 9) that integrates the data presented here with 354

current knowledge derived from previous studies in the cereal seed. 355

356

HvVP1 effects during barley seed development 357

In Hordeum vulgare, several TFs such as HvGAMYB, BPBF and the 358

bZIPs BLZ1 and BLZ2 (Vicente-Carbajosa et al., 1997; Mena et al., 1998; 359

Vicente-Carbajosa et al., 1998; Oñate et al., 1999; Díaz et al., 2002), acting as 360

part of regulatory complexes have been reported to participate in the regulation 361

of maturation genes like SSPs (B-hordein; gene HvHor2). Among them, the 362

HvGAMYB protein activates transcription of the HvHor2 gene by binding to the 363

corresponding MYB-R2R3 box (5´-TAACAAC-3´) in its promoter. Moreover, 364

HvGAMYB physically interacts with BPBF (Díaz et al., 2002) and together 365

significantly increase the trans-activation of the HvHor2 promoter in developing 366

barley endosperms as compared with the transactivation of each TF considered 367

separately. 368

In this work we have performed a detailed expression analysis of HvVP1 during 369

seed development to define its precise expression pattern. Since HvVP1 is a 370

known regulator of seed maturation genes, we wanted to survey its potential 371

effect on key regulatory factors controlling this process. As a first step, we 372

explored HvVP1 action on the transactivation of the HvHor2 promoter mediated 373

by HvGAMYB and demonstrate that HvVP1 interferes with its activation 374



16

capacity (Fig. 4B). Furthermore, our work shows that HvVP1 and HvGAMYB 375

interact in vivo both in yeast (Y2H) and in plant nuclei, in support that a direct 376

HvVP1-GAMYB protein-protein interaction takes place. Thus, we considered 377

whether the observed negative effects on the HvGAMYB-mediated 378

transactivation are derived from the HvVP1-HvGAMYB interaction.. 379

Interestingly, our results show that HvVP1 interferes with the HvGAMYB-BPBF 380

interaction (Y3H assays; Fig. 7 A & B), and also reduces the binding affinity of 381

both HvGAMYB and BPBF for their corresponding cis- motives in the HvHor2 382

promoter (Fig. 7C). However, as reflected in our proposed model (Fig 9) this 383

repressing activity of HvVP1 is not displayed on maturation genes (e.g. Hor2) in 384

the developing starchy endosperm, due to a non-overlapping temporal pattern 385

of expression. Notably, HvVP1 transcript levels decrease as maturation 386

progresses while the expression of HvGAMYB and BPBF increases, hindering 387

the possibility of HvGAMYB being kidnapped by HvVP1 and rendering a fully 388

operative activation complex. 389

Besides the HvVP1 expression pattern in the starchy endorsperm, its 390

persistence in the developing aleurone and immature embryos would favor the 391

repression of the α-amylase and other germination related genes, necessary for 392

acquisition of primary dormancy and for avoidance of pre-harvest sprouting, as 393

has been previously documented for its orthologs in maize and Arabidopsis 394

(Parcy et al., 1994; Hoecker et al., 1995). 395

396

HvVP1 effects during barley germination and post-germination phases 397



17

After root protrusion (post-germination), different genes encoding 398

hydrolytic enzymes, such as α-amylases, proteases, etc. that mobilize storage 399

reserves, are expressed in the aleurone. In the promoters of these genes a 400

conserved tripartite cis-acting element, the GA-Responsive Complex (GARC), 401

has been described to contain a GA-Responsive Element (GARE, 5’-402

TAACAAA-3’), the pyrimidine box (5’-CCTTTT-3’) and the 5’-TATCCAC-3’ box 403

(Sun and Gubler, 2004). In barley, the interaction of these three cis-elements 404

with TFs belonging to the MYBR2R3 (HvGAMYB), DOF (BPBF and three 405

others) and MYBR1-SHAQKYF (HvMCB1, HvMYBS3) families has been 406

demonstrated (Gubler, et al. 1999; Mena et al. 2002; Isabel-Lamoneda et al. 407

2003; Rubio-Somoza et al. 2006a, b; Moreno-Risueño et al. 2007). HvGAMYB 408

is considered the master regulator of hydrolase gene expression, and is highly 409

induced by GA in the aleurone of germinating seeds (Gubler et al. 1999, 2002). 410

In agreement with our observations during seed maturation, our results show 411

that HvVP1 also interferes with the HvGAMYB trans-activation of the Amy6.4 412

promoter in germinating aleurone cells (Fig. 4C). Again, this effect is sustained 413

by our disclosure that the interaction with HvVP1 leads to a decreased affinity of 414

GAMYB and other TFs (SAD- HvDOF23) to their DNA targets, as well as to the 415

interference in their protein-complex formation. Accordingly, HvGAMYB-416

dependent activation of post-germination genes can only be achieved once 417

HvVP1 expression disappears from the germinating aleurone. 418

As reflected in our model (Fig. 9), our results show that upon seed 419

imbibition HvVP1 is highly expressed in the mature embryo and in the aleurone 420

up to 24 hoi (the time at which root protrusion occurs), then sharply decaying, in 421

contrast to HvGAMYB whose expression rises up to 48 hoi (Diaz et al., 2002). 422



18

Taken together, these data indicate that HvVP1 not only prevents premature 423

expression of post-germination genes (encoding α-amylases, proteases, etc.), 424

but also that its disappearance during post-germination is required for 425

mobilization of reserves to take place. The transcriptional repression of 426

hydrolase genes mediated by VP1/ABI3 during germination has been previously 427

reported in monocot- and dicot- seeds, in species like maize, tomato and 428

Arabidopsis (Hoecker et al., 1999; Nambara et al., 2000; Bassel et al., 2006). 429

430

431

In conclusion, the precise spatio-temporal expression of HvVP1 allows 432

the fine-tune regulation of maturation and germination programs both within 433

distinct seed compartments and at different developmental stages. As disclosed 434

here, HvVP1 action occurs mainly by modulating the DNA-binding capacity and 435

protein-protein interactions of transcription factors, like HvGAMYB and BPBF 436

that participate in different regulatory complexes. 437

The HvVP1 effects described here rely mostly on its repressing activity or 438

the alternative activation derived from its release. In addition, an intrinsic 439

activation capacity has also been reported for the VP1/ABI3 proteins. In 440

particular, in non-endospermic seeds, commonly found in dicot plants like 441

Arabidopsis thaliana, ABI3 has been described as an activator of maturation 442

(SSP) and LEA genes, as reflected by their impaired expression in abi3 mutants 443

(Parcy et al., 1994). Moreover, ectopic over-expression of ABI3 (35S::ABI3) in 444

conjunction with bZIP factors (AtbZIP10, AtbZIP25, AtbZIP53) induces some of 445

the SSP genes in vegetative tissues, in support that ABI3 is a positive regulator 446

of SSP gene expression (Lara et al., 2003; Alonso et al., 2009). In this respect, 447



19

the observation that HvVP1 functions as a transcriptional repressor of the 448

HvHor2 promoter upon seed maturation is not contradictory, since in 449

Arabidopsis SSP and LEA proteins are expressed in the embryo, in contrast to 450

their accumulation in the cereal endosperm. Moreover, globulin storage proteins 451

similar to those of non-endospermic seeds are accumulated in the embryos of 452

cereal seeds and display a similar positive regulation under VP1, in support of a 453

conserved function of VP1/ABI3 in the control of SSP gene expression. Hence, 454

the molecular mechanisms underlying the regulatory properties of VP1/ABI3 455

proteins seem to be preserved, but differentially displayed in the context of 456

endosperm and embryo where interactions with a different set of trasnscription 457

factors occur. 458

459

460

461

462

MATERIALS AND METHODS 463

Plant Material and Growth Conditions 464

Barley (Hordeum vulgare) cv. Bomi seeds were germinated in the dark, 465

stratified at 4°C for four days, and then, grown in the greenhouse. Developing 466

endosperms (5, 10, 15 and 20 days after pollination; dap) and germinating 467

seeds (16, 24, 48 and 72 h of imbibition; hoi) were frozen in liquid N2 and stored 468

at -80 °C until used for RNA extraction. 469

Developing endosperms from cv. Bomi (15 dap) and aleurone layers 470

peeled off from barley cv. Himalaya germinating seeds (48 hoi) were collected 471

for particle bombardment experiments, and used immediately. 472



20

473

Screening of a Barley Genomic Library 474

A Lambda phage FIX II genomic library from 8 days old barley seedlings 475

(cv. Igri; Agilent Technologies), representing 16 x 108 plaque forming units 476

(pfu)/ml, was plated after infection of the Escherichia coli strain XL1-Blue MRA 477

(Agilent Technologies). The plaques were transferred onto nylon membranes 478

that were hybridized with a specific probe of 465-bp fragment obtained by PCR 479

with primers derived from conserved sequences of previously described VP1 480

genes in other cereals (Genebank references: Avena fatua: AJ001140; Oryza 481

sativa cv. Japonica, D16640.1; Sorghum bicolor, AF249881.1; Triticum 482

aestivum, AB047554.1; Zea mays, M60214.1; for primers see Supplemental 483

Table S2). This fragment spanned the B3 domain and a 3´-end conserved 484

region, and was α-[32P]-labeled, following standard procedures. Pre-485

hybridization, hybridization and membrane washing were done as described 486

(Vicente-Carbajosa et al., 1998). The in vivo excision properties of the Lambda 487

phage FIX II vector system have allowed the recovery of selected clones in the 488

pBluescript SK plasmid for sequencing (Agilent Technologies). 489

The HvVP1 ORF was obtained by PCR from cDNA isolated from 490

developing barley seeds (20 dap; see Supplemental Table S2), and its 491

corresponding deduced protein appears in Supplemental Fig. S1. 492

493

Bioinformatic Resources: HvVP1 Identification, Phylogenetic Dendrogram 494

and Transcriptomic Analysis 495

Identification of the HvVP1 genomic sequence (AJ431703.1) and its 496

corresponding deduced protein (CAD24413.1) was done using the bioinformatic 497



21

tools at the European Molecular Biology Laboratory (EMBL) and the National 498

Centre for Biotechnology Information (NCBI) databases. The Interpro Program 499

(PFAM database; Bateman et al., 2002; http://pfam.sanger.ac.uk) was used to 500

confirm the presence of the B3 DNA-binding domain. 501

The deduced protein sequences of VP1 genes were obtained from public 502

databases: those of Hordeum vulgare cv. Morex (International Barley Genome 503

Sequencing Consortium, 2012), and those from cvs Himalaya and Haruna Nijo 504

(HvVP1), Triticum aestivum (TaVP1), Triticum turgidum (TtVP1) and Triticum 505

monococcum (TmVP1) from Genbank (http://www.ncbi.nlm.nih.gov/genbank/); 506

and those of Brachypodium distachyon (BdVP1), Oryza sativa (OsVP1) and 507

Arabidopsis thaliana (AtABI3) from Phytozome v8.0 Database (Goodstein et al., 508

2012; www.phytozome.net). These sequences were aligned by means of the 509

CLUSTAL W program (Thompson et al., 1994) and utilized to construct a 510

phylogenetic dendrogram, using the neighbour-joining algorithm, a bootstrap 511

analysis with 1,000 replicates, complete deletion and the Jones Taylor Thornton 512

matrix, as settings. The MEME program software version 6.0 (Tamura et al., 513

2013) was used to identify conserved motives within the deduced VP1 proteins 514

and to validate the phylogenetic tree (Table 1). Default parameters were used, 515

except for: the maximum number of motives to find that was set to 21 and the 516

minimum width to 6 amino-acid residues (Bailey et al., 2009; 517

http://meme.sdsc.edu/meme4_6_0/intro.htlm). A single capital letter is given 518

when the frequency of a residue is greater than twice that of the second most 519

frequent one at the same position. A pair of capital letters in brackets represent 520

two residues with a relative frequency sum of >75%. When these criteria are not 521



22

satisfied, a lower-case letter is set when the relative frequency of a residue is 522

>40%, if not, x is given. 523

The major biochemical parameters of the deduced VP1 proteins are 524

listed in Supplemental Table S1. Both Isoeletric points (Ip) and Molecular 525

Weights (MW) were predicted using the “Compute pI/MW” tool (Gasteiger et al., 526

2005; http://www.expasy.ch/tools/pi_tool.html). 527

The in silico transcriptomic analysis of genes HvVP1, HvGAMYB, BPBF 528

and BLZ2 in Hordeum vulgare cv. Morex (Supplemetal Fig. S4) was done using 529

the Hordeum eFP browser at the Bio-Analytic Resource for Plant Biology 530

(http://bar.utoronto.ca/efpbarley/cgi-bin/efpWeb.cgi; Druka et al., 2006). For 531

identification of the probe sets for HvVP1, HvGAMYB, BPBF and BLZ2 532

transcription analysis the PLEXdb Blast was used 533

(http://www.plexdb.org/modules/tools/plexdb_blast.php). 534

Total RNA Isolation and Northern Blot Analysis 535

Total RNA was isolated from 1-2 g of barley developing de-embrionated 536

seeds (endosperms at 5, 10, 15, 20 dap), immature (20 dap) and mature dry 537

embryos, germinating aleurones and germinating embryos (8, 16, 24, 48, 72 538

hoi). After electrophoresis and transfer to nylon membranes, hybridization was 539

carried out at 68 °C, following standard procedures (Moreno-Risueño et al., 540

2007). A 280 bp fragment at the 3´-UTR (nt 3528-3808) was used as a specific 541

HvVP1 probe (see Supplemental Table S2). The 18S-RNA gene was used as a 542

control for sample charge. 543

544



23

mRNA In Situ Hybridization Experiments 545

The protocol described here is a modification of that described in Lara et 546

al. (2003). Barley developing (20 dap) and germinating seeds (24 hoi) were 547

collected and fixed in 4% para-formaldehyde (w/v), embedded in paraffin, 548

sectioned to 8 µm and de-waxed. An antisense or sense digoxigenin (DIG)-549

labelled RNA probes, corresponding to the same DNA fragment (280 bp) used 550

for Northern blots (see Supplemental Table S2), were synthesized with the DIG 551

RNA labelling mix and hybridization was at 56ºC overnight. Antibody incubation 552

and colour detection was done according to the manufacturer’s instructions 553

(Boehringer Ingelheim). 554

555

Yeast 1-, 2- and 3-Hybrid Assays 556

For yeast 2-Hybrid (Y2H) assays, the effector plasmids pGBT9 and 557

pGAD424 (Clontech), which contain the alcohol dehydrogenase I (AdhI) 558

promoter fused to the Gal4 DNA binding domain (Gal4BD; pGBT9 bait vector) 559

or to the Gal4 DNA activation domain (Gal4AD; pGAD424 prey vector), 560

respectively, were used to generate translational fusions with HvVP1, 561

HvGAMYB, BPBF ORFs or with selected fragments derived from HvVP1 that 562

were cloned into the EcoRI-BamHI sites by a PCR strategy. The haploid strains 563

SFY526 and HF7c of Saccharomyces cerevisiae (Clontech), carrying the LacZ 564

(β-galactosidase) and HIS3 (imidazole glycerol phosphate dehydratase) 565

reporter genes, under the control of a truncated Gal1 promoter that contains 566

Gal4-responsive elements (Gal1UAS), were used. 567



24

For yeast 3-Hybrid (Y3H) assays, the BPBF and the HvVP1 ORFs were 568

cloned into the pBridgeTM vector (Clontech) in the Multiple Cloning Sites I and II 569

(MCSI and MCSII), respectively. The BPBF ORF was cloned into the EcoRI-570

BamHI sites of MSCI fused to the GAL4 binding domain, and that of HvVP1 into 571

the NotI-BglII sites (MSCII) by a PCR strategy (see Table S2 for primer 572

sequences). Gal4AD-HvGAMYB (pGAD424 vector) previously used for Y2H 573

assays was used as the third component. The pBridgeTM vector allows the 574

conditional expression of a second gene under the control of the MET25 575

promoter (PMET25) in response to methionine (Met) levels in the medium. 576

All the Y2H and Y3H assays were done following the manufacture´s 577

instructions (Clontech). 578

579

580

Bimolecular Fluorescence Complementation (BiFC) 581

Essentially, HvVP1 and HvGAMYB ORFs were translationally fused to 582

the N- or to the C-terminal encoding fragments of the Green Fluorescent Protein 583

(GFP) gene ORF by using a PCR strategy and subsequent subcloning into the 584

two versions of the psmRS-GFP plasmid (psmRS-N-GFP or psmRS-C-GFP). 585

Final constructs were used to co-bombard inner epidermal cell layers of fresh 586

onions (Allium cepa) using a biolistic Helium gun device (DuPont PDS-1000; 587

BioRad Laboratories), as previously described (Díaz et al., 2005). The 588

fluorescence emission was observed after 36 h of incubation at 22ºC in the 589

dark, with a Carl Zeiss Axiophot fluorescence microscope (filter parameters: 590

excitation 450-490 nm; emission 520 nm). As a control of nuclei localization, the 591

onion cell layers were stained with DAPI (Serva). 592



25

Transient Expression Assays in Barley Developing Endosperms and in 593

germinating Aleurone Layers 594

The reporter vectors, containing the promoters of genes Hor2 (encoding 595

a B-hordein) and Amy6.4 (encoding a high-Ip barley α-amylase), PHor2 and 596

PAmy6.4, fused to the GUS reporter gene were previously described (Mena et al., 597

1998, 2002). The effector constructs corresponding to HvVP1, HvGAMYB and 598

BPBF, were prepared by cloning the corresponding ORFs in the pMF6 plasmid 599

under the control of the 35S CaMV promoter followed by the first intron of the 600

maize AdhI gene (35S-I); using a PCR strategy, followed by subcloning at the 601

EcoRI-BamHI sites of this plasmid. Particle bombardment and subsequent GUS 602

evaluation were carried out with a biolistic helium gun device (DuPont PSD-603

1000) as previously described (Mena et al., 1998, 2002; Díaz et al., 2002). 604

605

Electrophoretic Mobility Shift Assays (EMSAs) 606

Translational fusions of HvVP1, HvGAMYB, BPBF, SAD and BLZ2 ORFs 607

to the glutathione S-transferase (GST) gene were prepared by cloning their 608

ORFs into the expression vector pGEX-2T (Pharmacia). To this purpose, a PCR 609

strategy was done followed by the sub-cloning of these ORFs into the BamHI-610

EcoRI sites of the pGEX-2T plasmid (see Supplemental Table S2). Induction of 611

recombinant proteins with 0.1 mM IPTG (Isopropyl-β-d-Thio-Galato-Pyranoside) 612

and preparation of the E. coli protein extracts were performed as previously 613

described (Vicente-Carbajosa et al., 1997). 614

Two probes containing, one of them, the binding sites for HvVP1 besides 615

those for BLZ2 and BPBF (PB/GLM, see Fig. 7C) and the other that for 616

GAMYB (5´-TAACAAC-3´) within the Hor-2 promoter (PHor2), were produced by 617



26

annealing complementary single-stranded oligonucleotides that generate 618

protruding ends (see Supplemental Table S2). These probes were end-labeled 619

with [32P]-dATP by the fill-in reaction (Klenow exo-free DNA polymerase; United 620

States Biochemical) and purified from an 8% polyacrylamide gel electrophoresis 621

(39:1 cross-linking). The DNA-protein binding reactions and the EMSA 622

experiments were done, as described previously, using 1 ng of [32P]-labeled 623

probes (Moreno-Risueño et al., 2008). 624

625

ACKNOWLEDGMENTS 626

We thank Mar González for excellent technical assistance. 627

628

SUPPLEMENTAL DATA 629

Supplemental Fig. S1. Structure of the HvVP1 gene derived from cv. Igri. 630

Predicted intron-exon boundaries were confirmed by sequencing of the 631

corresponding cDNA; splicing signals are underlined. The deduced encoded 632

protein sequence is shown (in blue). 633

Supplemental Fig. S2. Gene copy number of HvVP1. Total DNA from barley 634

leaves was digested with restriction endonucleases: BamHI (B), EcoRI (E), SacI 635

(S), XbaI (X) and hybridized with a specific probe derived from 3´-non coding 636

region of the gene. 637

Supplemental Fig. S3. Comparison of the VP1 deduced sequences from 638

Hordeum vulgare cv. Igri (HvVP1), cv Morex (MLOC_69727), cv Haruna Nijo 639

(BAK06992.1), and a partial sequence from cv Himalaya (AAO06117.1). 640

Supplemental Fig. S4. Pictographic representation of HvVP1, GAMYB, BPBF 641

and BLZ2 transcript accumulation during seed development (5 to 22 dap) by 642

using the barley eFP browser of the Bio-Analytic Resource for Plant Biology 643

(http://bar.utoronto.ca/efpbarley/cgi-bin/efpWeb.cgi). 644



27

Supplemental Fig. S5. Transactivation assays using as effectors HvVP1 and 645

BPBF, and as reporter the Amy6.4 gene promoter driving the expression of the 646

uidA gene in the presence of gibberellins (GA). 647

Supplemental Fig. S6. Electrophoretic Mobility Shift Assays (EMSA) of the 648

recombinant proteins BLZ2 (bZIP family) and SAD (DOF family) with the 649

PB/GLM probe derived from the Hor2 gene promoter containing the cis-650

interacting motives for SAD and BLZ2 in the presence or absence of HvVP1. 651

Supplemental Table S1. Major characteristics of VP1/ABI3 predicted proteins. 652

Suppemental Table S2. Oligonucleotide sequences of primers used. 653

654

655



28

LEGENDS TO FIGURES 656

Figure 1. A, Phylogenetic tree with the deduced amino acid sequences of the 657

VP1 genes from distinct cvs of Hordeum vulgare (HvVP1 Igri, Morex and 658

Haruna Nijo) , Triticum aestivum (TaVP1), Triticum turgidum (TtVP1), Triticum 659

monococum (TmVP1), Brachypodium distachyon (BdVP1) and Oryza sativa 660

(OsVP1), and with AtABI3 from Arabidopsis thaliana; bootstrapping values are 661

specified in the branches. B, Distribution of the conserved motives among the 662

deduced protein sequences in the dendrogram (A), found by means of the 663

MEME analysis. A, B1, B2, and B3 correspond to conserved domains, as 664

described by Nakamura and Toyama (2001) and Marella and Quatrano (2007). 665

666

Figure 2. A, HvVP1 transcript splicing model. Exons I to VI are indicated as 667

solid bars. Motives A, B1, B2 and B3, as in Figure 1. B, Functional assay in 668

yeast for the activation capacity of the HvVP1 protein. Different constructs (1-6) 669

are generated as fusions to the Gal4 DNA-binding domain (black box at the N-670

terminus) and the activation capacity assayed in two alternative reporter 671

systems: ß-Galactosidase activity (LacZ reporter gene; Miller´s Units in liquid 672

medium), and HIS3 (growth capacity in Histidine depleted medium). 673

674

Figure 3. HvVP1 expression analyses by Northern Blot and by mRNA in situ 675

hybridization analysis, in barley developing (A) and germinating seeds (B). A.1 676

Northern blot analyses, 8 μg of total RNA from developing endosperms (from 5, 677

10, 15, 20 dap) and immature (25 dap) and mature embryos has been loaded in 678

each lane and hybridized to an HvVP1 gene-specific probe. A.2 HvVP1 679

transcript localization in longitudinal sections of developing (25 dap) seeds by 680

mRNA in situ hybridization B.1 Northern blot analysis in germinating aleurone 681

(16, 24, 48, 72 hoi) and germinating embryo (8, 16, 24 hoi). B.2 HvVP1 682

transcript localization in germinating barley seeds (36 hoi), by mRNA in situ 683

hybridization assays. Left and right panels correspond to hybridizations with 684

HvVP1 antisense and sense probes, respectively. The EtBr-stained rRNA 685

picture is included as a loading control. e: endosperm, em: embryo; p: pericarp; 686

s: scutellum. 687



29

Figure 4. Trans-activation assays using as effectors the TFs HvVP1 and 688

HvGAMYB. The Hor2 and Amy6.4 gene promoters driving the expression of the 689

uidA gene (GUS activity) were used as reporters. A, Schematic representation 690

of the effector and reporter constructs used in the analyses. B, Co-691

bombardment in barley developing endosperms of the effector and reporter 692

combinations indicated. C, The effector and reporter constructs designated 693

were co-bombarded in germinating aleurones from barley kernels. The relative 694

amounts of reporter and effector plasmids used in these assays correspond to 695

the 1:1 ratio. Values are the means ± SE of three independent replicates. 696

697

Figure 5. Protein-protein interaction by yeast two-hybrid assays. A, The 698

structures of HvGAMYB and HvVP1 are depicted showing the location of 699

functionally relevant domains within the proteins. B, HvGAMYB (complete ORF) 700

or HvVP1 N464 (the 464 N-terminal aminoacid residues) expressed as 701

translational fusions to the Gal4 DNA-binding (BD) and Activation Domains 702

(AD), respectively. C, Interaction capacity assayed by measuring the ß-703

Galactosidase activity (LacZ reporter gene; Miller Units in liquid medium) and in 704

D, by evaluating the yeast growth capacity in a Histidine depleted medium 705

(reporter gene: HIS3) with increasing concentrations of 3-aminotriazole (3-AT). 706

707

Figure 6. In planta interaction of HvVP1 and HvGAMYB. A, Structure of gene 708

constructs used in Bimolecular Fluorescence Complementation (BiFC) assays 709

using particle gun transformation of onion (Allium cepa) epidermal cells. B, 710

Green Fluorescent Protein (GFP) reconstruction in the nuclei of cells 711

transformed with a combination of the constructs described above (b). Bright 712

field images (a) and 4’,6-diamidino-2-phenylindole (DAPI) staining used to 713

reveal nuclei within the cells (c). 714

Figure 7. Protein-protein interactions of HvVP1, GAMYB and BPBF in yeast 715

3-hybrid experiments (Y3H) and the effect of HvVP1 on GAMYB and BPBF 716

DNA-binging capacities. A, Schematic representation of the constructs used in 717

the Y3H assays, where the BPBF-ORF was translationally fused to the Gal4 718



30

DNA binding domain (BD) in the pBridgeTM plasmid harboring HvVP1 under a 719

Methionine (Met) repressible promoter. The GAMYB-ORF was translationally 720

fused to the Gal4 activation domain (AD) in pGAD424. B, Schematic 721

representation of the constructs used for the Y3H assays and quantification of 722

β-galactosidase activity (expressed as Miller’s units) in liquid assays using for 723

the constructs represented above. The proteins expressed in Met-depleted 724

medium are indicated at the right side of the panel. C, EMSAs of HvGAMYB 725

and BPBF proteins in the absence (-) or presence (+) of the HvVP1 protein. 726

Incubations were done with a specific 32P-labeled probe containing a target 727

site for the corresponding transcription factors. MBS: MYB Binding Site 728

(GAMYB), PB: Prolamin Box (BPBF) , GLM: G-Box Like-Motif (BLZ2). 729

730

Figure 8. Transactivation assays using, as effectors, the TFs HvVP1 and BPBF 731

and, as reporter, the Hor2 gene promoter driving the expression of the uidA 732

gene (GUS activity). A, Schematic representation of the effector and the 733

reporter constructs used in the analyses. B, Co-bombardment experiments on 734

barley developing endosperms using the indicated combination of effector and 735

reporter constructs. The relative amount of reporter to effector plasmids used in 736

these assays corresponds to the 1:1 ratio. Values are means ± SE of three 737

independent replicates. 738

739

Figure 9. Proposed model of transcriptional regulation of SSP (Hor2) and 740

hydrolase (Amy6.4) genes in barley seeds mediated by HvVP1 interacting with 741

GAMYB, DOF (BPBF, SAD), BLZ2 and MR1 (MYBR1) TFs, during the 742

maturation and post-germination phases. The starchy endosperm and the 743

embryo plus aleurone layer are considered separately. Relative gene 744

expression levels are indicated in the y-axis. The dotted vertical line indicates 745

the time of root emergence. 746

747

748

749



31

Table 1 Sequences of conserved amino acid motives (MEME; Bailey et al. 750 2009) of the VIVIPAROUS-1 orthologous genes from Hordeum vulgare, 751 Triticum monococcum, Triticum turgidum, Triticum aestivum, Oryza sativa, 752 Brachypodium distachyon and Arabidopsis thaliana. 753

RKKR: Nuclear Localization Signal underlined in bold. 754 755

756

757

758

Motif E-value Consensus sequences

1 4.0e-377 DIGTS[QR]VW[SN]MRYRFWPNNKSRMYLLENTGDFVRSNELQEGDFIV[LI]YSDVK 2 8.8e-310 NNR[DE]CISAEDLRSIRL[RK]RSTIEAAAARLGGGRQGTMQLLKLILTWVQNHH 3 3.3e-175 DDFMFA[QD]DTFPALPDFPCLSSPSSS[TN]FSSSSSSNSSSAF 4 1.1e-150 DKNLRFLLQKVLKQSDVG[TS]LGRIVLPK[KE][EA] 5 3.6e-109 MEP[AS]AT[RK]EARKKRMARQRRLS[CS] 6 1.6e-097 EPSEPAAAGDG[MV]DDL[SA]DID[HQ]LLDFAS[IL][NS] 7 1.1e-149 YEFP[TA]ETGAAAATSWMPYQAFSPT[AG]SYGGEA[MI]YPFQQGCST 8 1.4e-151 DMHAGAWPLQYAAFVPAGATSAGTQTYPMPPPG[AP]VPQPFAAPGFAGQFPQ 9 5.4e-101 DDVPWDDEPLFPDVGMMLEDVISEQQ[QL]QQ

10 5.7e-101 KYLIRGVKVRA[AQ]Q[EG]LAKHKN[AG]SPEKGGAS[DE]VKA 11 2.1e-112 LQQQRSQQLNLSQIQTGGFPQE[PQ]SPRAAHSAPV

12 1.5e-123 [HG]W[SG][PG][PL][AW][VT]Q[AQ][QA][PV][HQ]GQLM[IV]QVPNPLSTKSNSSRQKQQKPSPDAAARPPSGGA

13 5.3e-087 EDGGCKEKSPHGVRRSRQEA[AS] 14 5.0e-080 ETHLPELKT[GR]DGISI 15 2.4e-071 GGG[GS][STG]GSAADDLP[RAL]FFMEWL[TK] 16 8.2e-074 LQKKRPRVGAMDQEAPPAGGQLPSPGANP 17 4.4e-042 SSV[VA]VSSQPFSPPAA 18 8.1e-042 [AH]P[QR][GR][GS][GA]P[HR]RGK[GA]PAVEIR[HQ]GE 19 1.2e-036 [HY]G[AT][AG]GR[TV]AS[DH]AA[AG]GGGEDAFM 20 2.6e-029 [ST][PQ][QH]R[PQ]GQA[AS]AS[DN]KQRQQGA[RS][TR][PT]AAAP[AP]AG 21 1.6e-026 MNQMAVSI



32

LITERATURE CITED 759

Alonso R, Oñate-Sánchez L, Weltmeier F, Ehlert A, Diaz I, Dietrich K, 760

Vicente-Carbajosa J, Dröge-Laser W (2009) A pivotal role of the basic 761

leucine zipper transcription factor bZIP53 in the regulation of Arabidopsis 762

seed maturation gene expression based on heterodimerization and protein 763

complex formation. Plant Cell 21:1747-61 764

Bailey TL, Boden M, Buske FA, Frith M, Grant CE, Clementi L, Ren J, Li 765

WW, Noble WS (2009) MEME SUITE: tools for motifs discovery and 766

searching. Nucleic Acid Res 37: W202–W208. 767

Bassel GW, Mullen RT, Bewley JD (2006) ABI3 expression ceases following, 768

but not during, germination of tomato and Arabidopsis seeds. J Exp Bot 769

57: 1291–1297 770

Bateman A, Birney E, Cerruti L, Durbin R, Etwiller L, Eddy SR, Griffiths-771

Jones S, Howe KL, Marshall M, Sonnhammer EL (2002) The Pfam 772

protein families database. Nucleic Acid Res 30: 276–280. 773

Bewley JD (1997) Seed germination and dormancy. Plant Cell 9: 1055–1066 774

Braybrook SA, Harada JJ (2008) LECs go crazy in embryo development. 775

Trends Plant Sci 13: 624–630 776

Casaretto J, Ho TH (2003) The transcription factors HvABI5 and HvVP1 are 777

required for the abscisic acid induction of gene expression in barley 778

aleurone cells. Plant Cell 15: 271–284 779

Diaz I, Martinez M, Isabel-LaMoneda I, Rubio-Somoza I, Carbonero P (2005) 780

The DOF protein, SAD, interacts with GAMYB in plant nuclei and activates 781

transcription of endosperm-specific genes during barley seed 782

development. Plant J 42: 652–662 783

Diaz I, Vicente-Carbajosa J, Abraham Z, Martínez M, Isabel-La Moneda I, 784

Carbonero P (2002) The GAMYB protein from barley interacts with the 785



33

DOF transcription factor BPBF and activates endosperm-specific genes 786

during seed development. Plant J 29: 453–464 787

Druka A, Muehlbauer G, Druka I, Caldo R, Baumann U, Rostoks N, 788

Schreiber A, Wise R, Close T, Kleinhofs A, Graner A, Schulman A, 789

Langridge P, Sato K, Hayes P, McNicol J, Marshall D, Waugh R (2006) 790

An atlas of gene expression from seed to seed through barley 791

development. Funct Integr Genomics 6: 202–211 792

Finch-Savage WE, Leubner-Metzger G (2006) Seed dormancy and the control 793

of germination. New Phytol 171: 501–523 794

Finkelstein RR, Gampala SS, Rock CD (2002) Abscisic acid signaling in 795

seeds and seedlings. Plant Cell 4 Suppl: S15–S45 796

Finkelstein R, Reeves W, Ariizumi T, Steber C (2008) Molecular aspects of 797

seed dormancy. Annu Rev Plant Biol 59: 387–415 798

Gasteiger E, Hoogland C, Gattiker A, Duvaud S, Wilkins MR, Appel RD, 799

Bairoch A (2005) Protein identification and analysis tools on the ExPASy 800

server. In Walker JK ed, The proteomics protocols handbook. Humana 801

Press, New York, pp 571–607 802

González-Calle V, Barrero-Sicilia C, Carbonero P, Iglesias-Fernández R 803

(2015) Mannans and endo-β-mannanases (MAN) in Brachypodium 804

distachyon: expression profiling and possible role of the BdMAN genes 805

during coleorhiza-limited seed germination. J Exp Bot: erv168 806

González-Calle V, Iglesias-Fernández R, Carbonero P, Barrero-Sicilia C 807

(2014) The BdGAMYB protein from Brachypodium distachyon interacts 808

with BdDOF24 and regulates transcription of the BdCathB gene upon 809

seed germination. Planta 240: 539–552 810

Goodstein DM, Shu S, Howson R, Neupane R, Hayes RD, Fazo J, Mitros T, 811

Dirks W, Hellsten, U, Putnam N, Rokhsar DS (2012) Phytozome: a 812

comparative platform for green plant genomics. Nucleic Acids Res 40: 813

D1178–1186 814



34

Graeber K, Linkies A, Müller K, Wunchova A, Rott A, Leubner-Metzger G. 815

2010. Cross-species approaches to seed dormancy and germination: 816

conservation and biodiversity of ABA-regulated mechanisms and the 817

Brassicaceae DOG1 genes. Plant Mol Biol. 73: 67–87 818

Gubler F, Chandler PM, White RG, Llewellyn DJ, Jacobsen JV (2002) 819

Gibberellin signaling in barley aleurone cells. Control of SLN1 and GAMYB 820

expression. Plant Physiol 129: 191–200 821

Gubler F, Kalla R, Roberts JK, Jacobsen JV (1995) Gibberellin-regulated 822

expression of a myb gene in barley aleurone cells: evidence for Myb 823

transactivation of a high-pI alpha-amylase gene promoter. Plant Cell 7: 824

1879–1891 825

Gubler F, Raventos D, Keys M, Watts R, Mundy J, Jacobsen JV (1999) 826

Target genes and regulatory domains of the GAMYB transcriptional 827

activator in cereal aleurone. Plant J 17: 1–9 828

Gubler F, Millar AA, Jacobsen JV (2005) Dormancy release, ABA and pre-829

harvest sprouting. Curr Opin Plant Biol 8: 183–187 830

Hattori T, Vasil V, Rosenkrans L, Hannah LC, McCarty DR, Vasil IK (1992) 831

The Viviparous-1 gene and abscisic acid activate the C1 regulatory gene 832

for anthocyanin biosynthesis during seed maturation in maize. Genes Dev 833

6: 609–618 834

Hill A, Nantel A, Rock CD, Quatrano RS (1996) A conserved domain of the 835

Viviparous-1 gene product enhances the DNA binding activity of the bZIP 836

protein EmBP-1 and other transcription factors. J Biol Chem 27:3366-74 837

838

Hobo T, Kowyama Y, Hattori T (1999) A bZIP factor, TRAB1, interacts with 839

VP1 and mediates abscisic acid-induced transcription. Proc Natl Acad Sci 840

U S A. 96:15348-53 841

Hoecker U, Vasil IK, McCarty DR (1995) Integrated control of seed maturation 842

and germination programs by activator and repressor functions of 843

Viviparous-1 of maize. Genes Dev 9: 2459–2469 844



35

Hoecker U, Vasil IK, McCarty DR (1999) Signaling from the embryo conditions 845

Vp1-mediated repression of alpha-amylase genes in the aleurone of 846

developing maize seeds. Plant J 19: 371–377 847

Holdsworth MJ, Bentsink L, Soppe WJJ (2008) Molecular networks 848

regulating Arabidopsis seed maturation, after-ripening, dormancy and 849

germination. New Phytol 179: 33–54 850

Iglesias-Fernández R, Rodríguez-Gacio MC, Matilla AJ (2011a) Progress in 851

research on afterripening. Seed Sci Res 21: 69–80 852

Iglesias-Fernández R, Rodríguez-Gacio MC, Barrero-Sicilia C, Carbonero 853

P, Matilla AJ (2011b) Three endo-β-mannanase genes expressed in the 854

micropylar endosperm and in the radicle influence germination of 855

Arabidopsis thaliana seeds. Planta 233: 25–36 856

Iglesias-Fernández R, Rodríguez-Gacio MC, Barrero-Sicilia C, Carbonero 857

P, Matilla AJ (2011c) Molecular analysis of endo-β-mannanase genes 858

upon seed imbibition suggests a cross-talk between radicle and micropylar 859

endosperm during germination of Arabidopsis thaliana. Plant Signal Behav 860

6: 80–82 861

Iglesias-Fernández R, Wozny D, Iriondo-de Hond M, Oñate-Sánchez L, 862

Carbonero P, Barrero-Sicilia C (2014) The AtCathB3 gene, encoding a 863

cathepsin B-like protease, is expressed during germination of Arabidopsis 864

thaliana and transcriptionally repressed by the basic leucine zipper protein 865

GBF1. J Exp Bot 65: 2009–2021 866

International Barley Genome Sequencing Consortium, Mayer KF, Waugh 867

R, Brown JW, Schulman A, Langridge P, Platzer M, Fincher GB, 868

Muehlbauer GJ, Sato K, Close TJ, Wise RP, Stein N (2012) A physical, 869

genetic and functional sequence assembly of the barley genome. Nature 870

491: 711–716 871

Isabel-LaMoneda I, Diaz I, Martinez M, Mena M, Carbonero P (2003) SAD: a 872

new DOF protein from barley that activates transcription of a cathepsin 873



36

B-like thiol protease gene in the aleurone of germinating seeds. Plant J 874

33: 329–340 875

Jones HD, Peters NC, Holdsworth MJ (1997) Genotype and environment 876

interact to control dormancy and differential expression of the 877

VIVIPAROUS 1 homologue in embryos of Avena fatua. Plant J 12: 911–878

920 879

Koornneef M, Bentsink L, Hilhorst H (2002) Seed dormancy and germination. 880

Curr Opin Plant Biol 5: 33–36 881

Lara P, Oñate-Sánchez L, Abraham Z, Ferrándiz C, Díaz I, Carbonero P, 882

Vicente-Carbajosa J (2003) Synergistic activation of seed storage protein 883

gene expression in Arabidopsis by ABI3 and two bZIPs related to 884

OPAQUE2. J Biol Chem 278: 21003–21011 885

Marella HH, Quatrano RS (2007) The B2 domain of VIVIPAROUS1 is bi-886

functional and regulates nuclear localization and transactivation. Planta 887

225: 863–872 888

Matsumoto T, Tanaka T, Sakai H, Amano N, Kanamori H, Kurita K, Kikuta 889

A, Kamiya K, Yamamoto M, Ikawa H, Fujii N, Hori K, Itoh T, Sato K. 890

2011. Comprehensive sequence analysis of 24,783 barley full-length 891

cDNAs derived from 12 clone libraries. Plant Physiol. 156: 20–28 892

McCarty DR, Carson CB, Stinard PS, Robertson DS (1989) Molecular 893

analysis of viviparous-1: an abscisic acid-insensitive mutant of maize. 894

Plant Cell 1: 523–532 895

McCarty DR, Hattori T, Carson CB, Vasil V, Lazar M, Vasil IK (1991) The 896

Viviparous-1 developmental gene of maize encodes a novel transcriptional 897

activator. Cell 66: 895–905 898

Mena M, Cejudo FJ, Isabel-Lamoneda I, Carbonero P (2002) A role for the 899

DOF transcription factor BPBF in the regulation of gibberellin-responsive 900

genes in barley aleurone. Plant Physiol 130: 111–119 901



37

Mena M, Vicente-Carbajosa J, Schmidt RJ, Carbonero P (1998) An 902

endosperm-specific DOF protein from barley, highly conserved in wheat, 903

binds to and activates transcription from the prolamin-box of a native B-904

hordein promoter in barley endosperm. Plant J 16: 53–62 905

Moreno-Risueño MA, Díaz I, Carrillo L, Fuentes R, Carbonero P (2007) The 906

HvDOF19 transcription factor mediates the abscisic acid-dependent 907

repression of hydrolase genes in germinating barley aleurone. Plant J 51: 908

352–365 909

Moreno-Risueño MA, González N, Díaz I, Parcy F, Carbonero P, Vicente-910

Carbajosa J (2008) FUSCA3 from barley unveils a common 911

transcriptional regulation of seed-specific genes between cereals and 912

Arabidopsis. Plant J 53: 882–894 913

Nakamura S, Toyama T (2001) Isolation of a VP1 homologue from wheat and 914

analysis of its expression in embryos of dormant and non-dormant 915

cultivars. J Exp Bot 52: 875–876 916

Nambara E, Hayama R, Tsuchiya Y, Nishimura M, Kawaide H, Kamiya Y, 917

Naito S (2000) The role of ABI3 and FUS3 loci in Arabidopsis thaliana on 918

phase transition from late embryo development to germination. Dev Biol 919

220: 412–423 920

Nonogaki H, Chen F, Bradford KJ (2007) Mechanisms and genes involved in 921

germination sensu stricto. In Bradford K y Nonogaki H eds, Seed 922

development, dormancy and germination. Blackwell Publishing, Oxford, pp 923

264–304 924

Oñate L, Vicente-Carbajosa J, Lara P, Díaz I, Carbonero P (1999) Barley 925

BLZ2, a seed-specific bZIP protein that interacts with BLZ1 in vivo and 926

activates transcription from the GCN4-like motif of B-hordein promoters in 927

barley endosperm. J Biol Chem 274: 9175–9182 928

Parcy F, Valon C, Raynal M, Gaubier-Comella P, Delseny M, Giraudat J 929

(1994) Regulation of gene expression programs during Arabidopsis seed 930



38

development: roles of the ABI3 locus and of endogenous abscisic acid. 931

Plant Cell 6: 1567–1582 932

Paz-Ares J, Wienand U, Peterson PA, Saedler H. (1986) Molecular cloning of 933

the c locus of Zea mays: a locus regulating the anthocyanin pathway. 934

EMBO J 5: 829-833 935

Pritchard SL, Charlton WL, Baker A, Graham IA (2002) Germination and 936

storage reserve mobilization are regulated independently in Arabidopsis. 937

Plant J. 31: 639–647 938

Reidt W, Wohlfarth T, Ellerström M, Czihal A, Tewes A, Ezcurra I, Rask L, 939

Bäumlein H (2000) Gene regulation during late embryogenesis: the RY 940

motif of maturation-specific gene promoters is a direct target of the FUS3 941

gene product. Plant J 21: 401–408 942

Rodríguez MV, Barrero JM, Corbineau F, Gubler F, Benech-Arnold RL 943

(2015) Dormancy in cereals (not too much, not so little): about the 944

mechanisms behind this trait. Seed Sci Res 25: 99–119 945

Rubio-Somoza I, Martinez M, Abraham Z, Diaz I, Carbonero P (2006a) 946

Ternary complex formation between HvMYBS3 and other factors 947

involved in transcriptional control in barley seeds. Plant J 47: 269–281 948

Rubio-Somoza I, Martinez M, Diaz I, Carbonero P (2006b) HvMCB1, a 949

R1MYB transcription factor from barley with antagonistic regulatory 950

functions during seed development and germination. Plant J 45: 17–30 951

Sun TP, Gubler F. (2004) Molecular mechanism of gibberellin signaling in 952

plants. Annu Rev Plant Biol 55: 197-223. 953

Suzuki M, Kao CY, Cocciolone S, McCarty DR (2001) Maize VP1 954

complements Arabidopsis abi3 and confers a novel ABA/auxin 955

interaction in roots. Plant J 28:409–418. 956

Swaminathan K, Peterson K, Jack T (2008) The plant B3 superfamily. Trends 957

Plant Sci 13: 647–655 958



39

Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: 959

Molecular Evolutionary Genetics Analysis version 6.0. Mol Biol Evol 30: 960

2725–2729 961

Thompson J, Higgins D, Gibson T (1994) CLUSTAL W: improving the 962

sensitivity of progressive multiple sequence alignment through sequence 963

weighting, position-specific gap penalties and weight matrix choice. 964

Nucleic Acids Res 22: 4673–4680 965

To A, Valon C, Savino G, Guilleminot J, Devic M, Giraudat J, Parcy F 966

(2006) A network of local and redundant gene regulation governs 967

Arabidopsis seed maturation. Plant Cell 18: 1642–1651 968

Vasil V, Marcotte WR Jr, Rosenkrans L, Cocciolone SM, Vasil IK, Quatrano 969

RS, McCarty DR (1995) Overlap of Viviparous1 (VP1) and abscisic acid 970

response elements in the Em promoter: G-box elements are sufficient but 971

not necessary for VP1 transactivation. Plant Cell 7: 1511–1508 972

Vicente-Carbajosa J, Carbonero P (2005) Seed maturation: developing an 973

intrusive phase to accomplish a quiescent state. Int J Dev Biol 49: 645–974

651 975

Vicente-Carbajosa J, Moose SP, Parsons RL, Schmidt RJ (1997) A maize 976

zinc-finger protein binds the prolamin box in zein gene promoters and 977

interacts with the basic leucine zipper transcriptional activator Opaque2. 978

Proc Natl Acad Sci USA 94: 7685–7690 979

Vicente-Carbajosa J, Oñate L, Lara P, Diaz I, Carbonero P (1998) Barley 980

BLZ1: a bZIP transcriptional activator that interacts with endosperm-981

specific gene promoters. Plant J 13: 629–640 982

Yan D, Duermeyer L, Leoveanu C, Nambar E (2014) The functions of the 983

endosperm during seed germination. Plant Cell Physiol 55: 1521-1533 984



40

Zeng Y, Raimondi N, Kermode AR (2003) Role of an ABI3 homologue in 985

dormancy maintenance of yellow-cedar seeds and in the activation of 986

storage protein and Em gene promoters. Plant Mol Biol 51: 39–49 987

Zeng Y, Zhao T, Kermode AR (2013) A conifer ABI3-interacting protein plays 988

important roles during key transitions of the plant life cycle. Plant Physiol 989

161: 179–195 990



Figure1

Figure1.A,Phylogene,ctreewiththededucedaminoacidsequencesoftheVP1genesfromdis,nctcvsofHordeumvulgare(HvVP1Igri,MorexandHarunaNijo),Tri8cumaes8vum(TaVP1),Tri8cumturgidum(TtVP1),Tri8cummonococum(TmVP1),Brachypodiumdistachyon(BdVP1)andOryzasa8va(OsVP1),andwithAtABI3fromArabidopsisthaliana;bootstrappingvaluesarespecifiedinthebranches.B,Distribu,onoftheconservedmo,vesamongthededucedprotein

sequencesinthedendrogram(A),foundbymeansoftheMEMEanalysis.A,B1,B2,

andB3correspondtoconserveddomains,asdescribedbyNakamuraandToyama

(2001)andMarellaandQuatrano(2007).

HvVP1(Igri) 18 3 6 9 19 15 2 16 7 17 8 5 11 12 20 4 14 1 10 13 21HvVP1(Morex) 18 3 6 9 19 15 2 16 7 17 8 5 11 12 20 4 14 1 10 13 21HvVP1 (Haruna) 18 3 6 9 19 15 2 16 7 17 8 5 11 12 20 4 14 1 10 13 21

TmVP1 18 3 6 9 19 15 2 16 7 17 8 5 11 12 20 4 14 1 10 13 21TaVP1 18 3 6 9 19 15 2 16 7 17 8 5 11 12 20 4 14 1 10 13 21TtVP1 18 3 6 9 19 15 2 16 7 17 8 5 11 12 20 4 14 1 10 13 21OsVP1 18 3 6 15 2 7 17 5 11 4 14 1 13 21BdVP1 18 3 6 15 2 17 5 4 14 1 13 21AtABI3 3 15 2 5 4 14 1

A B1 B2 B3

A

B

90

100100

100

10099 HvVP1 (Igri)

HvVP1 (Morex)HvVP1 (Haruna Nijo)

TtVP1TaVP1

OsVP1BdVP1AtABI3

TriticeaeTmVP1



I II III IV V VI

A B1 B3B2

1 24- 127 182 - 375 381- 408 519- 629 682 bp

A

BA B1 B3B2BD

A B1 B2BD

ABD

B1 B2BD

B3BD

1

2

3

4

5

ß-Gal M.U. His-

12345

200 400 6000

Figure2.A,HvVP1transcriptsplicingmodel.ExonsItoVIareindicatedassolidbars.

Mo,ves A, B1, B2 and B3, as in Figure 1. B, Func,onal assay in yeast for the

ac,va,oncapacityoftheHvVP1protein.Differentconstructs(1-6)aregeneratedas

fusions to the Gal4 DNA-binding domain (black box at the N-terminus) and the

ac,va,on capacity assayed in two alterna,ve reporter systems: ß-Galactosidase

ac,vity (LacZ reporter gene; Miller´s Units in liquid medium), and HIS3 (growthcapacityinHis,dinedepletedmedium).

Figure2



Figure3.HvVP1expressionanalysesbyNorthernBlotandbymRNA insituhybridiza,onanalysis,inbarleydeveloping(A)andgermina,ngseeds(B).A.1Northernblotanalyses,8

μgoftotalRNAfromdevelopingendosperms(from5,10,15,20dap)andimmature(25

dap)andmatureembryoshasbeenloadedineachlaneandhybridizedtoanHvVP1gene-specificprobe.A.2HvVP1transcriptlocaliza,oninlongitudinalsec,onsofdeveloping(25dap) seeds by mRNA in situ hybridiza,on B.1 Northern blot analysis in germina,ng

aleurone(16,24,48,72hoi)andgermina,ngembryo(8,16,24hoi).B.2HvVP1transcriptlocaliza,oningermina,ngbarleyseeds(36hoi),bymRNAinsituhybridiza,onassays.Lecand right panels correspond to hybridiza,onswithHvVP1 an,sense and sense probes,respec,vely.TheEtBr-stainedrRNApictureisincludedasaloadingcontrol.e:endosperm,

em:embryo;p:pericarp;s:scutellum.

Figure3



Figure 4. Trans-ac,va,on assays using as effectors the TFsHvVP1 andHvGAMYB. The

Hor2andAmy6.4genepromotersdrivingtheexpressionoftheuidAgene(GUSac,vity)were used as reporters. A, Schema,c representa,on of the effector and reporter

constructsusedintheanalyses.B,Co-bombardmentinbarleydevelopingendospermsof

the effector and reporter combina,ons indicated. C, The effector and reporter

constructs designated were co-bombarded in germina,ng aleurones from barley

kernels. The rela,ve amounts of reporter and effector plasmids used in these assays

correspondtothe1:1ra,o.Valuesarethemeans±SEofthreeindependentreplicates

P35S

I-ADHI

A" Effectors

PAmy 6.4 (- 174 bp) uidA

PAmy 6.4 :uidA

100

400

900

Rel

GU

S A

ctiv

ity (%

)

PHor2 (- 560 bp)

Reporters

B"

100

300

500

HvVP1 HvGAMYB

1 2 3 4

Rel

GU

S A

ctiv

ity (%

) PHor2:uidA

C"

HvVP1

--

-+

+-

++

Seed Maturation

Seed Germination

HvVP1 HvGAMYB

1 2 3 4

--

-+

+-

++

HvGAMYB

Tnos

Tnos

Figure4



A

A B1 B3B2

1 24- 127 182- 375 381- 408 519- 629 682

A1BD

1 4- 150 151- 230 356- 490 553

A2

HvVP1HvGAMYB

AD / BD Constructs

Gal4 ADGal4 BD

HvVP1 N1 +

HvGAMYB2 +

HvGAMYB3

464

HvVP1 N464

100

200

300

400

1 2 3

β-G

al M

.U.

0 mM

60 mM

45 mM

30 mM

15 mM

AD-Ø[ 3-AT ] AD-HvVP1 N

BD-HvGAMYB

+

C

B

D

Figure5.Protein-protein interac,onbyyeast two-hybridassays.A,ThestructuresofHvGAMYBandHvVP1aredepictedshowingtheloca,onoffunc,onallyrelevant

domainswithintheproteins.B,HvGAMYB(completeORF)orHvVP1N464 (the464

N-terminalaminoacidresidues)expressedastransla,onalfusionstotheGal4DNA-

binding (BD) and Ac,va,on Domains (AD), respec,vely. C, Interac,on capacity

assayedbymeasuringtheß-Galactosidaseac,vity(LacZreportergene;MillerUnits

in liquidmedium)and inD,byevalua,ng theyeastgrowthcapacity inaHis,dine

depleted medium (reporter gene: HIS3) with increasing concentra,ons of 3-aminotriazole(3-AT).

Figure5



Figure 6. In planta interac,on of HvVP1 and HvGAMYB. A, Structure of gene

constructsused inBimolecularFluorescenceComplementa,on (BiFC)assaysusing

par,cle gun transforma,on of onion (Allium cepa) epidermal cells. B, Green

Fluorescent Protein (GFP) reconstruc,on in thenuclei of cells transformedwith a

combina,onoftheconstructsdescribedabove(b).Brightfieldimages(a)and4’,6-

diamidino-2-phenylindole(DAPI)stainingusedtorevealnucleiwithinthecells(c).

A

B

a

b

c

HvVP1

GAMYB

p35S

p35S nos

nos GFP-N

GFP-C

Figure6



Figure7.Protein-proteininterac,onsofHvVP1,GAMYBandBPBFinyeast3-hybridexperiments

(Y3H) and the effect of HvVP1 on GAMYB and BPBF DNA-binging capaci,es. A, Schema,c

representa,onoftheconstructsusedintheY3Hassays,wheretheBPBF-ORFwastransla,onally

fusedtotheGal4DNAbindingdomain(BD) inthepBridgeTMplasmidharboringHvVP1undera

Methionine (Met) repressiblepromoter. TheGAMYB-ORFwas transla,onally fused to theGal4

ac,va,ondomain(AD) inpGAD424.B,Schema,crepresenta,onoftheconstructsusedforthe

Y3H assays and quan,fica,on of β-galactosidase ac,vity (expressed asMiller’s units) in liquid

assays using for the constructs represented above. The proteins expressed in Met-depleted

mediumareindicatedattherightsideofthepanel.C,EMSAsofHvGAMYBandBPBFproteinsin

theabsence(-)orpresence(+)oftheHvVP1protein.Incuba,onsweredonewithaspecific32P-

labeled probe containing a target site for the corresponding transcrip,on factors. MBS: MYB

BindingSite(GAMYB),PB:ProlaminBox(BPBF),GLM:G-BoxLike-Mo,f(BLZ2).

A

1!

GAMYB2!

VP1!BPBF!4!

BPBF!3!

B

VP1!BPBF!

GAMYB

- Met

- Met

| | | |

ß-Gal M.U.!

Gal4 ADGal4 BD

CGAMYB

+ + +

BPBF

- - - + + + +- - - - HvVP1

GAMYB probe (1) PB/GLM probe (2)

(1): CATATCTTAACAACCCACACACGATT

(2): GTGACATGTAAAGTGAATAAGGTGAGTCATGCA

1 + 3!

1 + 4!

2 + 3!

2 + 4!

PB! GLM!

BPBF

BPBF + VP1

BPBF + GAMYB

BPBF + GAMYB + VP1

0! 20! 40! 60! 80!

MBS!

Figure7



Figure8.Transac,va,onassaysusing,aseffectors, theTFsHvVP1andBPBFand,as reporter,theHor2 genepromoterdriving theexpressionof theuidA gene (GUSac,vity).A, Schema,c

representa,on of the effector and the reporter constructs used in the analyses. B, Co-

bombardmentexperimentsonbarleydevelopingendospermsusingtheindicatedcombina,on

ofeffectorandreporterconstructs.Therela,veamountofreportertoeffectorplasmidsusedin

these assays corresponds to the 1:1 ra,o. Values are means ± SE of three independent

replicates.

P35S

I-ADHI

A Effectors

uidAPHor2 (- 560 bp)

Reporters

B

150

450

750

HvVP1 BPBF

1 2 3 4

Rel

GU

S A

ctiv

ity (%

) PHor2 :uidA

BPBF

--

-+

+-

++

Seed Maturation

Tnos

HvVP1 Tnos

Figure8



Figure 9.Proposedmodel of transcrip1onal regula1on of SSP (Hor2) and hydrolase (Amy6.4)genesinbarleyseedsmediatedbyHvVP1interac1ngwithGAMYB,DOF(BPBF,SAD),BLZ2andMR1(MYBR1)TFs,duringthematura1onandpost-germina1onphases.Thestarchyendospermandtheembryoplusaleuronelayerareconsideredseparately.Rela1vegeneexpressionlevelsareindicatedinthey-axis.ThedoPedver1callineindicatesthe1meofrootemergence.

Figure9

MATURATION GERMINATION & POST-GERM.

STAR

CHYEN

DOSPER

M

EMBR

YO&ALEURO

NELA

YER

HvVP1GAMYB

5 10 15 >20 16 24 48 72hoi

HvVP1

GAMYB

Hor22 Hor22x

Amy6.4x

PBF

SAD

BLZ2

MR1

Amy6.4

em

end

al

Parsed CitationsAlonso R, Oñate-Sánchez L, Weltmeier F, Ehlert A, Diaz I, Dietrich K, Vicente-Carbajosa J, Dröge-Laser W (2009) A pivotal role ofthe basic leucine zipper transcription factor bZIP53 in the regulation of Arabidopsis seed maturation gene expression based onheterodimerization and protein complex formation. Plant Cell 21:1747-61

Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Bailey TL, Boden M, Buske FA, Frith M, Grant CE, Clementi L, Ren J, Li WW, Noble WS (2009) MEME SUITE: tools for motifsdiscovery and searching. Nucleic Acid Res 37: W202-W208.


Bassel GW, Mullen RT, Bewley JD (2006) ABI3 expression ceases following, but not during, germination of tomato and Arabidopsisseeds. J Exp Bot 57: 1291-1297


Bateman A, Birney E, Cerruti L, Durbin R, Etwiller L, Eddy SR, Griffiths-Jones S, Howe KL, Marshall M, Sonnhammer EL (2002) ThePfam protein families database. Nucleic Acid Res 30: 276-280.


Bewley JD (1997) Seed germination and dormancy. Plant Cell 9: 1055-1066Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Braybrook SA, Harada JJ (2008) LECs go crazy in embryo development. Trends Plant Sci 13: 624-630Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Casaretto J, Ho TH (2003) The transcription factors HvABI5 and HvVP1 are required for the abscisic acid induction of geneexpression in barley aleurone cells. Plant Cell 15: 271-284


Diaz I, Martinez M, Isabel-LaMoneda I, Rubio-Somoza I, Carbonero P (2005) The DOF protein, SAD, interacts with GAMYB in plantnuclei and activates transcription of endosperm-specific genes during barley seed development. Plant J 42: 652-662


Diaz I, Vicente-Carbajosa J, Abraham Z, Martínez M, Isabel-La Moneda I, Carbonero P (2002) The GAMYB protein from barleyinteracts with the DOF transcription factor BPBF and activates endosperm-specific genes during seed development. Plant J 29:453-464


Druka A, Muehlbauer G, Druka I, Caldo R, Baumann U, Rostoks N, Schreiber A, Wise R, Close T, Kleinhofs A, Graner A, SchulmanA, Langridge P, Sato K, Hayes P, McNicol J, Marshall D, Waugh R (2006) An atlas of gene expression from seed to seed throughbarley development. Funct Integr Genomics 6: 202-211


Finch-Savage WE, Leubner-Metzger G (2006) Seed dormancy and the control of germination. New Phytol 171: 501-523Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Finkelstein RR, Gampala SS, Rock CD (2002) Abscisic acid signaling in seeds and seedlings. Plant Cell 4 Suppl: S15-S45Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Finkelstein R, Reeves W, Ariizumi T, Steber C (2008) Molecular aspects of seed dormancy. Annu Rev Plant Biol 59: 387-415Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Gasteiger E, Hoogland C, Gattiker A, Duvaud S, Wilkins MR, Appel RD, Bairoch A (2005) Protein identification and analysis tools onthe ExPASy server. In Walker JK ed, The proteomics protocols handbook. Humana Press, New York, pp 571-607

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 21%5BVolume%5D AND 1747%5BPagination%5D AND Alonso R%2C O�ate%2DS�nchez L%2C Weltmeier F%2C Ehlert A%2C Diaz I%2C Dietrich K%2C Vicente%2DCarbajosa J%2C Dr�ge%2DLaser W%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Alonso R, O�ate-S�nchez L, Weltmeier F, Ehlert A, Diaz I, Dietrich K, Vicente-Carbajosa J, Dr�ge-Laser W (2009) A pivotal role of the basic leucine zipper transcription factor bZIP53 in the regulation of Arabidopsis seed maturation gene expression based on heterodimerization and protein complex formation. Plant Cell 21:1747-61

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Alonso&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=A pivotal role of the basic leucine zipper transcription factor bZIP53 in the regulation of Arabidopsis seed maturation gene expression based on heterodimerization and protein complex formation&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A pivotal role of the basic leucine zipper transcription factor bZIP53 in the regulation of Arabidopsis seed maturation gene expression based on heterodimerization and protein complex formation&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Alonso&as_ylo=2009&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Nucleic Acid Res%5BTitle%5D%29 AND 37%5BVolume%5D AND W202%5BPagination%5D AND Bailey TL%2C Boden M%2C Buske FA%2C Frith M%2C Grant CE%2C Clementi L%2C Ren J%2C Li WW%2C Noble WS%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Bailey TL, Boden M, Buske FA, Frith M, Grant CE, Clementi L, Ren J, Li WW, Noble WS (2009) MEME SUITE: tools for motifs discovery and searching. Nucleic Acid Res 37: W202-W208.

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Bailey&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=MEME SUITE: tools for motifs discovery and searching&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=MEME SUITE: tools for motifs discovery and searching&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Bailey&as_ylo=2009&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28J Exp Bot%5BTitle%5D%29 AND 57%5BVolume%5D AND 1291%5BPagination%5D AND Bassel GW%2C Mullen RT%2C Bewley JD%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Bassel GW, Mullen RT, Bewley JD (2006) ABI3 expression ceases following, but not during, germination of tomato and Arabidopsis seeds. J Exp Bot 57: 1291-1297

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Bassel&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=ABI3 expression ceases following, but not during, germination of tomato and Arabidopsis seeds&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=ABI3 expression ceases following, but not during, germination of tomato and Arabidopsis seeds&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Bassel&as_ylo=2006&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Nucleic Acid Res%5BTitle%5D%29 AND 30%5BVolume%5D AND 276%5BPagination%5D AND Bateman A%2C Birney E%2C Cerruti L%2C Durbin R%2C Etwiller L%2C Eddy SR%2C Griffiths%2DJones S%2C Howe KL%2C Marshall M%2C Sonnhammer EL%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Bateman A, Birney E, Cerruti L, Durbin R, Etwiller L, Eddy SR, Griffiths-Jones S, Howe KL, Marshall M, Sonnhammer EL (2002) The Pfam protein families database. Nucleic Acid Res 30: 276-280.

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Bateman&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The Pfam protein families database&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The Pfam protein families database&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Bateman&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 9%5BVolume%5D AND 1055%5BPagination%5D AND Bewley JD%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Bewley JD (1997) Seed germination and dormancy. Plant Cell 9: 1055-1066

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Bewley&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Seed germination and dormancy&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Seed germination and dormancy&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Bewley&as_ylo=1997&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Trends Plant Sci%5BTitle%5D%29 AND 13%5BVolume%5D AND 624%5BPagination%5D AND Braybrook SA%2C Harada JJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Braybrook SA, Harada JJ (2008) LECs go crazy in embryo development. Trends Plant Sci 13: 624-630

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Braybrook&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=LECs go crazy in embryo development&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=LECs go crazy in embryo development&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Braybrook&as_ylo=2008&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 15%5BVolume%5D AND 271%5BPagination%5D AND Casaretto J%2C Ho TH%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Casaretto J, Ho TH (2003) The transcription factors HvABI5 and HvVP1 are required for the abscisic acid induction of gene expression in barley aleurone cells. Plant Cell 15: 271-284

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Casaretto&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The transcription factors HvABI5 and HvVP1 are required for the abscisic acid induction of gene expression in barley aleurone cells&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The transcription factors HvABI5 and HvVP1 are required for the abscisic acid induction of gene expression in barley aleurone cells&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Casaretto&as_ylo=2003&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 42%5BVolume%5D AND 652%5BPagination%5D AND Diaz I%2C Martinez M%2C Isabel%2DLaMoneda I%2C Rubio%2DSomoza I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Diaz I, Martinez M, Isabel-LaMoneda I, Rubio-Somoza I, Carbonero P (2005) The DOF protein, SAD, interacts with GAMYB in plant nuclei and activates transcription of endosperm-specific genes during barley seed development. Plant J 42: 652-662

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Diaz&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The DOF protein, SAD, interacts with GAMYB in plant nuclei and activates transcription of endosperm-specific genes during barley seed development&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The DOF protein, SAD, interacts with GAMYB in plant nuclei and activates transcription of endosperm-specific genes during barley seed development&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Diaz&as_ylo=2005&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 29%5BVolume%5D AND 453%5BPagination%5D AND Diaz I%2C Vicente%2DCarbajosa J%2C Abraham Z%2C Mart�nez M%2C Isabel%2DLa Moneda I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Diaz I, Vicente-Carbajosa J, Abraham Z, Mart�nez M, Isabel-La Moneda I, Carbonero P (2002) The GAMYB protein from barley interacts with the DOF transcription factor BPBF and activates endosperm-specific genes during seed development. Plant J 29: 453-464

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Diaz&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The GAMYB protein from barley interacts with the DOF transcription factor BPBF and activates endosperm-specific genes during seed development&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The GAMYB protein from barley interacts with the DOF transcription factor BPBF and activates endosperm-specific genes during seed development&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Diaz&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Funct Integr Genomics%5BTitle%5D%29 AND 6%5BVolume%5D AND 202%5BPagination%5D AND Druka A%2C Muehlbauer G%2C Druka I%2C Caldo R%2C Baumann U%2C Rostoks N%2C Schreiber A%2C Wise R%2C Close T%2C Kleinhofs A%2C Graner A%2C Schulman A%2C Langridge P%2C Sato K%2C Hayes P%2C McNicol J%2C Marshall D%2C Waugh R%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Druka A, Muehlbauer G, Druka I, Caldo R, Baumann U, Rostoks N, Schreiber A, Wise R, Close T, Kleinhofs A, Graner A, Schulman A, Langridge P, Sato K, Hayes P, McNicol J, Marshall D, Waugh R (2006) An atlas of gene expression from seed to seed through barley development. Funct Integr Genomics 6: 202-211

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Druka&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=An atlas of gene expression from seed to seed through barley development&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=An atlas of gene expression from seed to seed through barley development&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Druka&as_ylo=2006&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28New Phytol%5BTitle%5D%29 AND 171%5BVolume%5D AND 501%5BPagination%5D AND Finch%2DSavage WE%2C Leubner%2DMetzger G%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Finch-Savage WE, Leubner-Metzger G (2006) Seed dormancy and the control of germination. New Phytol 171: 501-523

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Finch-Savage&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Seed dormancy and the control of germination&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Seed dormancy and the control of germination&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Finch-Savage&as_ylo=2006&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Abscisic acid signaling in seeds and seedlings%2E%5BTitle%5D%29 AND Finkelstein RR%2C Gampala SS%2C Rock CD%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Finkelstein RR, Gampala SS, Rock CD (2002) Abscisic acid signaling in seeds and seedlings. Plant Cell 4 Suppl: S15-S45

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Finkelstein&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Abscisic acid signaling in seeds and seedlings.&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Abscisic acid signaling in seeds and seedlings.&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Finkelstein&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Annu Rev Plant Biol%5BTitle%5D%29 AND 59%5BVolume%5D AND 387%5BPagination%5D AND Finkelstein R%2C Reeves W%2C Ariizumi T%2C Steber C%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Finkelstein R, Reeves W, Ariizumi T, Steber C (2008) Molecular aspects of seed dormancy. Annu Rev Plant Biol 59: 387-415

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Finkelstein&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular aspects of seed dormancy&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular aspects of seed dormancy&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Finkelstein&as_ylo=2008&as_allsubj=all&hl=en&c2coff=1


González-Calle V, Barrero-Sicilia C, Carbonero P, Iglesias-Fernández R (2015) Mannans and endo-ß-mannanases (MAN) inBrachypodium distachyon: expression profiling and possible role of the BdMAN genes during coleorhiza-limited seed germination.J Exp Bot: erv168


González-Calle V, Iglesias-Fernández R, Carbonero P, Barrero-Sicilia C (2014) The BdGAMYB protein from Brachypodiumdistachyon interacts with BdDOF24 and regulates transcription of the BdCathB gene upon seed germination. Planta 240: 539-552


Goodstein DM, Shu S, Howson R, Neupane R, Hayes RD, Fazo J, Mitros T, Dirks W, Hellsten, U, Putnam N, Rokhsar DS (2012)Phytozome: a comparative platform for green plant genomics. Nucleic Acids Res 40: D1178-1186


Graeber K, Linkies A, Müller K, Wunchova A, Rott A, Leubner-Metzger G. 2010. Cross-species approaches to seed dormancy andgermination: conservation and biodiversity of ABA-regulated mechanisms and the Brassicaceae DOG1 genes. Plant Mol Biol. 73:67-87


Gubler F, Chandler PM, White RG, Llewellyn DJ, Jacobsen JV (2002) Gibberellin signaling in barley aleurone cells. Control of SLN1and GAMYB expression. Plant Physiol 129: 191-200


Gubler F, Kalla R, Roberts JK, Jacobsen JV (1995) Gibberellin-regulated expression of a myb gene in barley aleurone cells:evidence for Myb transactivation of a high-pI alpha-amylase gene promoter. Plant Cell 7: 1879-1891


Gubler F, Raventos D, Keys M, Watts R, Mundy J, Jacobsen JV (1999) Target genes and regulatory domains of the GAMYBtranscriptional activator in cereal aleurone. Plant J 17: 1-9


Gubler F, Millar AA, Jacobsen JV (2005) Dormancy release, ABA and pre-harvest sprouting. Curr Opin Plant Biol 8: 183-187Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Hattori T, Vasil V, Rosenkrans L, Hannah LC, McCarty DR, Vasil IK (1992) The Viviparous-1 gene and abscisic acid activate the C1regulatory gene for anthocyanin biosynthesis during seed maturation in maize. Genes Dev 6: 609-618


Hill A, Nantel A, Rock CD, Quatrano RS (1996) A conserved domain of the Viviparous-1 gene product enhances the DNA bindingactivity of the bZIP protein EmBP-1 and other transcription factors. J Biol Chem 27:3366-74


Hobo T, Kowyama Y, Hattori T (1999) A bZIP factor, TRAB1, interacts with VP1 and mediates abscisic acid-induced transcription.Proc Natl Acad Sci U S A. 96:15348-53


Hoecker U, Vasil IK, McCarty DR (1995) Integrated control of seed maturation and germination programs by activator andrepressor functions of Viviparous-1 of maize. Genes Dev 9: 2459-2469


Hoecker U, Vasil IK, McCarty DR (1999) Signaling from the embryo conditions Vp1-mediated repression of alpha-amylase genes inthe aleurone of developing maize seeds. Plant J 19: 371-377

Pubmed: Author and Title

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Protein identification and analysis tools on the ExPASy server%2E%5BTitle%5D%29 AND Gasteiger E%2C Hoogland C%2C Gattiker A%2C Duvaud S%2C Wilkins MR%2C Appel RD%2C Bairoch A%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gasteiger E, Hoogland C, Gattiker A, Duvaud S, Wilkins MR, Appel RD, Bairoch A (2005) Protein identification and analysis tools on the ExPASy server. In Walker JK ed, The proteomics protocols handbook. Humana Press, New York, pp 571-607

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Gasteiger&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Protein identification and analysis tools on the ExPASy server.&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Protein identification and analysis tools on the ExPASy server.&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gasteiger&as_ylo=2005&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Mannans and endo%2D�%2Dmannanases MAN in Brachypodium distachyon%3A expression profiling and possible role of the BdMAN genes during coleorhiza%2Dlimited seed germination%2E%5BTitle%5D%29 AND Gonz�lez%2DCalle V%2C Barrero%2DSicilia C%2C Carbonero P%2C Iglesias%2DFern�ndez R%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gonz�lez-Calle V, Barrero-Sicilia C, Carbonero P, Iglesias-Fern�ndez R (2015) Mannans and endo-�-mannanases (MAN) in Brachypodium distachyon: expression profiling and possible role of the BdMAN genes during coleorhiza-limited seed germination. J Exp Bot: erv168

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Gonz�lez-Calle&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Mannans and endo-�-mannanases (MAN) in Brachypodium distachyon: expression profiling and possible role of the BdMAN genes during coleorhiza-limited seed germination.&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Mannans and endo-�-mannanases (MAN) in Brachypodium distachyon: expression profiling and possible role of the BdMAN genes during coleorhiza-limited seed germination.&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gonz�lez-Calle&as_ylo=2015&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Planta%5BTitle%5D%29 AND 240%5BVolume%5D AND 539%5BPagination%5D AND Gonz�lez%2DCalle V%2C Iglesias%2DFern�ndez R%2C Carbonero P%2C Barrero%2DSicilia C%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gonz�lez-Calle V, Iglesias-Fern�ndez R, Carbonero P, Barrero-Sicilia C (2014) The BdGAMYB protein from Brachypodium distachyon interacts with BdDOF24 and regulates transcription of the BdCathB gene upon seed germination. Planta 240: 539-552

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Gonz�lez-Calle&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The BdGAMYB protein from Brachypodium distachyon interacts with BdDOF24 and regulates transcription of the BdCathB gene upon seed germination&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The BdGAMYB protein from Brachypodium distachyon interacts with BdDOF24 and regulates transcription of the BdCathB gene upon seed germination&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gonz�lez-Calle&as_ylo=2014&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Nucleic Acids Res%5BTitle%5D%29 AND 40%5BVolume%5D AND D1178%5BPagination%5D AND Goodstein DM%2C Shu S%2C Howson R%2C Neupane R%2C Hayes RD%2C Fazo J%2C Mitros T%2C Dirks W%2C Hellsten%2C U%2C Putnam N%2C Rokhsar DS%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Goodstein DM, Shu S, Howson R, Neupane R, Hayes RD, Fazo J, Mitros T, Dirks W, Hellsten, U, Putnam N, Rokhsar DS (2012) Phytozome: a comparative platform for green plant genomics. Nucleic Acids Res 40: D1178-1186

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Goodstein&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Phytozome: a comparative platform for green plant genomics&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Phytozome: a comparative platform for green plant genomics&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Goodstein&as_ylo=2012&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Mol Biol%5BTitle%5D%29 AND 73%5BVolume%5D AND 67%5BPagination%5D AND Graeber K%2C Linkies A%2C M�ller K%2C Wunchova A%2C Rott A%2C Leubner%2DMetzger G%2E%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Graeber K, Linkies A, M�ller K, Wunchova A, Rott A, Leubner-Metzger G. 2010. Cross-species approaches to seed dormancy and germination: conservation and biodiversity of ABA-regulated mechanisms and the Brassicaceae DOG1 genes. Plant Mol Biol. 73: 67-87

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Graeber&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Cross-species approaches to seed dormancy and germination: conservation and biodiversity of ABA-regulated mechanisms and the Brassicaceae DOG1 genes&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Cross-species approaches to seed dormancy and germination: conservation and biodiversity of ABA-regulated mechanisms and the Brassicaceae DOG1 genes&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Graeber&as_ylo=2010&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Control of SLN1 and GAMYB expression%2E Plant Physiol%5BTitle%5D%29 AND 129%5BVolume%5D AND 191%5BPagination%5D AND Gubler F%2C Chandler PM%2C White RG%2C Llewellyn DJ%2C Jacobsen JV%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gubler F, Chandler PM, White RG, Llewellyn DJ, Jacobsen JV (2002) Gibberellin signaling in barley aleurone cells. Control of SLN1 and GAMYB expression. Plant Physiol 129: 191-200

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Gubler&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Gibberellin signaling in barley aleurone cells&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Gibberellin signaling in barley aleurone cells&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gubler&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 7%5BVolume%5D AND 1879%5BPagination%5D AND Gubler F%2C Kalla R%2C Roberts JK%2C Jacobsen JV%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gubler F, Kalla R, Roberts JK, Jacobsen JV (1995) Gibberellin-regulated expression of a myb gene in barley aleurone cells: evidence for Myb transactivation of a high-pI alpha-amylase gene promoter. Plant Cell 7: 1879-1891


http://scholar.google.com/scholar?as_q=Gibberellin-regulated expression of a myb gene in barley aleurone cells: evidence for Myb transactivation of a high-pI alpha-amylase gene promoter&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Gibberellin-regulated expression of a myb gene in barley aleurone cells: evidence for Myb transactivation of a high-pI alpha-amylase gene promoter&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gubler&as_ylo=1995&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 17%5BVolume%5D AND 1%5BPagination%5D AND Gubler F%2C Raventos D%2C Keys M%2C Watts R%2C Mundy J%2C Jacobsen JV%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gubler F, Raventos D, Keys M, Watts R, Mundy J, Jacobsen JV (1999) Target genes and regulatory domains of the GAMYB transcriptional activator in cereal aleurone. Plant J 17: 1-9


http://scholar.google.com/scholar?as_q=Target genes and regulatory domains of the GAMYB transcriptional activator in cereal aleurone&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Target genes and regulatory domains of the GAMYB transcriptional activator in cereal aleurone&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gubler&as_ylo=1999&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Curr Opin Plant Biol%5BTitle%5D%29 AND 8%5BVolume%5D AND 183%5BPagination%5D AND Gubler F%2C Millar AA%2C Jacobsen JV%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Gubler F, Millar AA, Jacobsen JV (2005) Dormancy release, ABA and pre-harvest sprouting. Curr Opin Plant Biol 8: 183-187


http://scholar.google.com/scholar?as_q=Dormancy release, ABA and pre-harvest sprouting&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Dormancy release, ABA and pre-harvest sprouting&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Gubler&as_ylo=2005&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Genes Dev%5BTitle%5D%29 AND 6%5BVolume%5D AND 609%5BPagination%5D AND Hattori T%2C Vasil V%2C Rosenkrans L%2C Hannah LC%2C McCarty DR%2C Vasil IK%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Hattori T, Vasil V, Rosenkrans L, Hannah LC, McCarty DR, Vasil IK (1992) The Viviparous-1 gene and abscisic acid activate the C1 regulatory gene for anthocyanin biosynthesis during seed maturation in maize. Genes Dev 6: 609-618

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Hattori&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The Viviparous-1 gene and abscisic acid activate the C1 regulatory gene for anthocyanin biosynthesis during seed maturation in maize&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The Viviparous-1 gene and abscisic acid activate the C1 regulatory gene for anthocyanin biosynthesis during seed maturation in maize&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Hattori&as_ylo=1992&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28J Biol Chem%5BTitle%5D%29 AND 27%5BVolume%5D AND 3366%5BPagination%5D AND Hill A%2C Nantel A%2C Rock CD%2C Quatrano RS%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Hill A, Nantel A, Rock CD, Quatrano RS (1996) A conserved domain of the Viviparous-1 gene product enhances the DNA binding activity of the bZIP protein EmBP-1 and other transcription factors. J Biol Chem 27:3366-74

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Hill&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=A conserved domain of the Viviparous-1 gene product enhances the DNA binding activity of the bZIP protein EmBP-1 and other transcription factors&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A conserved domain of the Viviparous-1 gene product enhances the DNA binding activity of the bZIP protein EmBP-1 and other transcription factors&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Hill&as_ylo=1996&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28A%5BTitle%5D%29 AND 1%5BVolume%5D AND interacts with VP1 and mediates abscisic acid%5BPagination%5D AND Hobo T%2C Kowyama Y%2C Hattori T%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Hobo T, Kowyama Y, Hattori T (1999) A bZIP factor, TRAB1, interacts with VP1 and mediates abscisic acid-induced transcription. Proc Natl Acad Sci U S A. 96:15348-53

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Hobo&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Hobo&as_ylo=1999&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Genes Dev%5BTitle%5D%29 AND 9%5BVolume%5D AND 2459%5BPagination%5D AND Hoecker U%2C Vasil IK%2C McCarty DR%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Hoecker U, Vasil IK, McCarty DR (1995) Integrated control of seed maturation and germination programs by activator and repressor functions of Viviparous-1 of maize. Genes Dev 9: 2459-2469

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Hoecker&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Integrated control of seed maturation and germination programs by activator and repressor functions of Viviparous-1 of maize&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Integrated control of seed maturation and germination programs by activator and repressor functions of Viviparous-1 of maize&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Hoecker&as_ylo=1995&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 19%5BVolume%5D AND 371%5BPagination%5D AND Hoecker U%2C Vasil IK%2C McCarty DR%5BAuthor%5D&dopt=abstract

CrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Holdsworth MJ, Bentsink L, Soppe WJJ (2008) Molecular networks regulating Arabidopsis seed maturation, after-ripening,dormancy and germination. New Phytol 179: 33-54


Iglesias-Fernández R, Rodríguez-Gacio MC, Matilla AJ (2011a) Progress in research on afterripening. Seed Sci Res 21: 69-80Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Iglesias-Fernández R, Rodríguez-Gacio MC, Barrero-Sicilia C, Carbonero P, Matilla AJ (2011b) Three endo-ß-mannanase genesexpressed in the micropylar endosperm and in the radicle influence germination of Arabidopsis thaliana seeds. Planta 233: 25-36


Iglesias-Fernández R, Rodríguez-Gacio MC, Barrero-Sicilia C, Carbonero P, Matilla AJ (2011c) Molecular analysis of endo-ß-mannanase genes upon seed imbibition suggests a cross-talk between radicle and micropylar endosperm during germination ofArabidopsis thaliana. Plant Signal Behav 6: 80-82


Iglesias-Fernández R, Wozny D, Iriondo-de Hond M, Oñate-Sánchez L, Carbonero P, Barrero-Sicilia C (2014) The AtCathB3 gene,encoding a cathepsin B-like protease, is expressed during germination of Arabidopsis thaliana and transcriptionally repressed bythe basic leucine zipper protein GBF1. J Exp Bot 65: 2009-2021


International Barley Genome Sequencing Consortium, Mayer KF, Waugh R, Brown JW, Schulman A, Langridge P, Platzer M,Fincher GB, Muehlbauer GJ, Sato K, Close TJ, Wise RP, Stein N (2012) A physical, genetic and functional sequence assembly ofthe barley genome. Nature 491: 711-716


Isabel-LaMoneda I, Diaz I, Martinez M, Mena M, Carbonero P (2003) SAD: a new DOF protein from barley that activatestranscription of a cathepsin B-like thiol protease gene in the aleurone of germinating seeds. Plant J 33: 329-340


Jones HD, Peters NC, Holdsworth MJ (1997) Genotype and environment interact to control dormancy and differential expressionof the VIVIPAROUS 1 homologue in embryos of Avena fatua. Plant J 12: 911-920


Koornneef M, Bentsink L, Hilhorst H (2002) Seed dormancy and germination. Curr Opin Plant Biol 5: 33-36Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Lara P, Oñate-Sánchez L, Abraham Z, Ferrándiz C, Díaz I, Carbonero P, Vicente-Carbajosa J (2003) Synergistic activation of seedstorage protein gene expression in Arabidopsis by ABI3 and two bZIPs related to OPAQUE2. J Biol Chem 278: 21003-21011


Marella HH, Quatrano RS (2007) The B2 domain of VIVIPAROUS1 is bi-functional and regulates nuclear localization andtransactivation. Planta 225: 863-872


Matsumoto T, Tanaka T, Sakai H, Amano N, Kanamori H, Kurita K, Kikuta A, Kamiya K, Yamamoto M, Ikawa H, Fujii N, Hori K, Itoh T,Sato K. 2011. Comprehensive sequence analysis of 24,783 barley full-length cDNAs derived from 12 clone libraries. Plant Physiol.156: 20-28


McCarty DR, Carson CB, Stinard PS, Robertson DS (1989) Molecular analysis of viviparous-1: an abscisic acid-insensitive mutantof maize. Plant Cell 1: 523-532

Pubmed: Author and Title

http://search.crossref.org/?page=1&rows=2&q=Hoecker U, Vasil IK, McCarty DR (1999) Signaling from the embryo conditions Vp1-mediated repression of alpha-amylase genes in the aleurone of developing maize seeds. Plant J 19: 371-377

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Hoecker&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Signaling from the embryo conditions Vp1-mediated repression of alpha-amylase genes in the aleurone of developing maize seeds&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Signaling from the embryo conditions Vp1-mediated repression of alpha-amylase genes in the aleurone of developing maize seeds&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Hoecker&as_ylo=1999&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28New Phytol%5BTitle%5D%29 AND 179%5BVolume%5D AND 33%5BPagination%5D AND Holdsworth MJ%2C Bentsink L%2C Soppe WJJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Holdsworth MJ, Bentsink L, Soppe WJJ (2008) Molecular networks regulating Arabidopsis seed maturation, after-ripening, dormancy and germination. New Phytol 179: 33-54

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Holdsworth&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular networks regulating Arabidopsis seed maturation, after-ripening, dormancy and germination&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular networks regulating Arabidopsis seed maturation, after-ripening, dormancy and germination&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Holdsworth&as_ylo=2008&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Seed Sci Res%5BTitle%5D%29 AND 21%5BVolume%5D AND 69%5BPagination%5D AND Iglesias%2DFern�ndez R%2C Rodr�guez%2DGacio MC%2C Matilla AJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Iglesias-Fern�ndez R, Rodr�guez-Gacio MC, Matilla AJ (2011a) Progress in research on afterripening. Seed Sci Res 21: 69-80

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Iglesias-Fern�ndez&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Progress in research on afterripening&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Progress in research on afterripening&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Iglesias-Fern�ndez&as_ylo=2011&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Planta%5BTitle%5D%29 AND 233%5BVolume%5D AND 25%5BPagination%5D AND Iglesias%2DFern�ndez R%2C Rodr�guez%2DGacio MC%2C Barrero%2DSicilia C%2C Carbonero P%2C Matilla AJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Iglesias-Fern�ndez R, Rodr�guez-Gacio MC, Barrero-Sicilia C, Carbonero P, Matilla AJ (2011b) Three endo-�-mannanase genes expressed in the micropylar endosperm and in the radicle influence germination of Arabidopsis thaliana seeds. Planta 233: 25-36


http://scholar.google.com/scholar?as_q=Three endo-�-mannanase genes expressed in the micropylar endosperm and in the radicle influence germination of Arabidopsis thaliana seeds&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Three endo-�-mannanase genes expressed in the micropylar endosperm and in the radicle influence germination of Arabidopsis thaliana seeds&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Iglesias-Fern�ndez&as_ylo=2011&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Signal Behav%5BTitle%5D%29 AND 6%5BVolume%5D AND 80%5BPagination%5D AND Iglesias%2DFern�ndez R%2C Rodr�guez%2DGacio MC%2C Barrero%2DSicilia C%2C Carbonero P%2C Matilla AJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Iglesias-Fern�ndez R, Rodr�guez-Gacio MC, Barrero-Sicilia C, Carbonero P, Matilla AJ (2011c) Molecular analysis of endo-�-mannanase genes upon seed imbibition suggests a cross-talk between radicle and micropylar endosperm during germination of Arabidopsis thaliana. Plant Signal Behav 6: 80-82


http://scholar.google.com/scholar?as_q=Molecular analysis of endo-�-mannanase genes upon seed imbibition suggests a cross-talk between radicle and micropylar endosperm during germination of Arabidopsis thaliana&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular analysis of endo-�-mannanase genes upon seed imbibition suggests a cross-talk between radicle and micropylar endosperm during germination of Arabidopsis thaliana&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Iglesias-Fern�ndez&as_ylo=2011&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28J Exp Bot%5BTitle%5D%29 AND 65%5BVolume%5D AND 2009%5BPagination%5D AND Iglesias%2DFern�ndez R%2C Wozny D%2C Iriondo%2Dde Hond M%2C O�ate%2DS�nchez L%2C Carbonero P%2C Barrero%2DSicilia C%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Iglesias-Fern�ndez R, Wozny D, Iriondo-de Hond M, O�ate-S�nchez L, Carbonero P, Barrero-Sicilia C (2014) The AtCathB3 gene, encoding a cathepsin B-like protease, is expressed during germination of Arabidopsis thaliana and transcriptionally repressed by the basic leucine zipper protein GBF1. J Exp Bot 65: 2009-2021


http://scholar.google.com/scholar?as_q=The AtCathB3 gene, encoding a cathepsin B-like protease, is expressed during germination of Arabidopsis thaliana and transcriptionally repressed by the basic leucine zipper protein GBF1&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The AtCathB3 gene, encoding a cathepsin B-like protease, is expressed during germination of Arabidopsis thaliana and transcriptionally repressed by the basic leucine zipper protein GBF1&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Iglesias-Fern�ndez&as_ylo=2014&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Nature%5BTitle%5D%29 AND 491%5BVolume%5D AND 711%5BPagination%5D AND International Barley Genome Sequencing Consortium%2C Mayer KF%2C Waugh R%2C Brown JW%2C Schulman A%2C Langridge P%2C Platzer M%2C Fincher GB%2C Muehlbauer GJ%2C Sato K%2C Close TJ%2C Wise RP%2C Stein N%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=International Barley Genome Sequencing Consortium, Mayer KF, Waugh R, Brown JW, Schulman A, Langridge P, Platzer M, Fincher GB, Muehlbauer GJ, Sato K, Close TJ, Wise RP, Stein N (2012) A physical, genetic and functional sequence assembly of the barley genome. Nature 491: 711-716

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=International&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=A physical, genetic and functional sequence assembly of the barley genome&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A physical, genetic and functional sequence assembly of the barley genome&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=International&as_ylo=2012&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 33%5BVolume%5D AND 329%5BPagination%5D AND Isabel%2DLaMoneda I%2C Diaz I%2C Martinez M%2C Mena M%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Isabel-LaMoneda I, Diaz I, Martinez M, Mena M, Carbonero P (2003) SAD: a new DOF protein from barley that activates transcription of a cathepsin B-like thiol protease gene in the aleurone of germinating seeds. Plant J 33: 329-340

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Isabel-LaMoneda&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=SAD: a new DOF protein from barley that activates transcription of a cathepsin B-like thiol protease gene in the aleurone of germinating seeds&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=SAD: a new DOF protein from barley that activates transcription of a cathepsin B-like thiol protease gene in the aleurone of germinating seeds&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Isabel-LaMoneda&as_ylo=2003&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 12%5BVolume%5D AND 911%5BPagination%5D AND Jones HD%2C Peters NC%2C Holdsworth MJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Jones HD, Peters NC, Holdsworth MJ (1997) Genotype and environment interact to control dormancy and differential expression of the VIVIPAROUS 1 homologue in embryos of Avena fatua. Plant J 12: 911-920

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Jones&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Genotype and environment interact to control dormancy and differential expression of the VIVIPAROUS 1 homologue in embryos of Avena fatua&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Genotype and environment interact to control dormancy and differential expression of the VIVIPAROUS 1 homologue in embryos of Avena fatua&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Jones&as_ylo=1997&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Curr Opin Plant Biol%5BTitle%5D%29 AND 5%5BVolume%5D AND 33%5BPagination%5D AND Koornneef M%2C Bentsink L%2C Hilhorst H%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Koornneef M, Bentsink L, Hilhorst H (2002) Seed dormancy and germination. Curr Opin Plant Biol 5: 33-36

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Koornneef&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Seed dormancy and germination&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Seed dormancy and germination&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Koornneef&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28J Biol Chem%5BTitle%5D%29 AND 278%5BVolume%5D AND 21003%5BPagination%5D AND Lara P%2C O�ate%2DS�nchez L%2C Abraham Z%2C Ferr�ndiz C%2C D�az I%2C Carbonero P%2C Vicente%2DCarbajosa J%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Lara P, O�ate-S�nchez L, Abraham Z, Ferr�ndiz C, D�az I, Carbonero P, Vicente-Carbajosa J (2003) Synergistic activation of seed storage protein gene expression in Arabidopsis by ABI3 and two bZIPs related to OPAQUE2. J Biol Chem 278: 21003-21011

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Lara&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Synergistic activation of seed storage protein gene expression in Arabidopsis by ABI3 and two bZIPs related to OPAQUE2&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Synergistic activation of seed storage protein gene expression in Arabidopsis by ABI3 and two bZIPs related to OPAQUE2&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Lara&as_ylo=2003&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Planta%5BTitle%5D%29 AND 225%5BVolume%5D AND 863%5BPagination%5D AND Marella HH%2C Quatrano RS%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Marella HH, Quatrano RS (2007) The B2 domain of VIVIPAROUS1 is bi-functional and regulates nuclear localization and transactivation. Planta 225: 863-872

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Marella&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The B2 domain of VIVIPAROUS1 is bi-functional and regulates nuclear localization and transactivation&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The B2 domain of VIVIPAROUS1 is bi-functional and regulates nuclear localization and transactivation&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Marella&as_ylo=2007&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Comprehensive%5BTitle%5D%29 AND 24%5BVolume%5D AND 783 barley full%5BPagination%5D AND Matsumoto T%2C Tanaka T%2C Sakai H%2C Amano N%2C Kanamori H%2C Kurita K%2C Kikuta A%2C Kamiya K%2C Yamamoto M%2C Ikawa H%2C Fujii N%2C Hori K%2C Itoh T%2C Sato K%2E%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Matsumoto T, Tanaka T, Sakai H, Amano N, Kanamori H, Kurita K, Kikuta A, Kamiya K, Yamamoto M, Ikawa H, Fujii N, Hori K, Itoh T, Sato K. 2011. Comprehensive sequence analysis of 24,783 barley full-length cDNAs derived from 12 clone libraries. Plant Physiol. 156: 20-28

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Matsumoto&hl=en&c2coff=1


http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Matsumoto&as_ylo=2011&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Molecular%5BTitle%5D%29 AND 1%5BVolume%5D AND McCarty DR%2C Carson CB%2C Stinard PS%2C Robertson DS%5BAuthor%5D&dopt=abstract

CrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

McCarty DR, Hattori T, Carson CB, Vasil V, Lazar M, Vasil IK (1991) The Viviparous-1 developmental gene of maize encodes anovel transcriptional activator. Cell 66: 895-905


Mena M, Cejudo FJ, Isabel-Lamoneda I, Carbonero P (2002) A role for the DOF transcription factor BPBF in the regulation ofgibberellin-responsive genes in barley aleurone. Plant Physiol 130: 111-119


Mena M, Vicente-Carbajosa J, Schmidt RJ, Carbonero P (1998) An endosperm-specific DOF protein from barley, highly conservedin wheat, binds to and activates transcription from the prolamin-box of a native B-hordein promoter in barley endosperm. Plant J16: 53-62


Moreno-Risueño MA, Díaz I, Carrillo L, Fuentes R, Carbonero P (2007) The HvDOF19 transcription factor mediates the abscisicacid-dependent repression of hydrolase genes in germinating barley aleurone. Plant J 51: 352-365


Moreno-Risueño MA, González N, Díaz I, Parcy F, Carbonero P, Vicente-Carbajosa J (2008) FUSCA3 from barley unveils a commontranscriptional regulation of seed-specific genes between cereals and Arabidopsis. Plant J 53: 882-894


Nakamura S, Toyama T (2001) Isolation of a VP1 homologue from wheat and analysis of its expression in embryos of dormant andnon-dormant cultivars. J Exp Bot 52: 875-876


Nambara E, Hayama R, Tsuchiya Y, Nishimura M, Kawaide H, Kamiya Y, Naito S (2000) The role of ABI3 and FUS3 loci in Arabidopsisthaliana on phase transition from late embryo development to germination. Dev Biol 220: 412-423


Nonogaki H, Chen F, Bradford KJ (2007) Mechanisms and genes involved in germination sensu stricto. In Bradford K y Nonogaki Heds, Seed development, dormancy and germination. Blackwell Publishing, Oxford, pp 264-304


Oñate L, Vicente-Carbajosa J, Lara P, Díaz I, Carbonero P (1999) Barley BLZ2, a seed-specific bZIP protein that interacts with BLZ1in vivo and activates transcription from the GCN4-like motif of B-hordein promoters in barley endosperm. J Biol Chem 274: 9175-9182


Parcy F, Valon C, Raynal M, Gaubier-Comella P, Delseny M, Giraudat J (1994) Regulation of gene expression programs duringArabidopsis seed development: roles of the ABI3 locus and of endogenous abscisic acid. Plant Cell 6: 1567-1582


Paz-Ares J, Wienand U, Peterson PA, Saedler H. (1986) Molecular cloning of the c locus of Zea mays: a locus regulating theanthocyanin pathway. EMBO J 5: 829-833


Pritchard SL, Charlton WL, Baker A, Graham IA (2002) Germination and storage reserve mobilization are regulated independentlyin Arabidopsis. Plant J. 31: 639-647


Reidt W, Wohlfarth T, Ellerström M, Czihal A, Tewes A, Ezcurra I, Rask L, Bäumlein H (2000) Gene regulation during lateembryogenesis: the RY motif of maturation-specific gene promoters is a direct target of the FUS3 gene product. Plant J 21: 401-408

http://search.crossref.org/?page=1&rows=2&q=McCarty DR, Carson CB, Stinard PS, Robertson DS (1989) Molecular analysis of viviparous-1: an abscisic acid-insensitive mutant of maize. Plant Cell 1: 523-532

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=McCarty&hl=en&c2coff=1


http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=McCarty&as_ylo=1989&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Cell%5BTitle%5D%29 AND 66%5BVolume%5D AND 895%5BPagination%5D AND McCarty DR%2C Hattori T%2C Carson CB%2C Vasil V%2C Lazar M%2C Vasil IK%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=McCarty DR, Hattori T, Carson CB, Vasil V, Lazar M, Vasil IK (1991) The Viviparous-1 developmental gene of maize encodes a novel transcriptional activator. Cell 66: 895-905

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=McCarty&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The Viviparous-1 developmental gene of maize encodes a novel transcriptional activator&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The Viviparous-1 developmental gene of maize encodes a novel transcriptional activator&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=McCarty&as_ylo=1991&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Physiol%5BTitle%5D%29 AND 130%5BVolume%5D AND 111%5BPagination%5D AND Mena M%2C Cejudo FJ%2C Isabel%2DLamoneda I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Mena M, Cejudo FJ, Isabel-Lamoneda I, Carbonero P (2002) A role for the DOF transcription factor BPBF in the regulation of gibberellin-responsive genes in barley aleurone. Plant Physiol 130: 111-119

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Mena&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=A role for the DOF transcription factor BPBF in the regulation of gibberellin-responsive genes in barley aleurone&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A role for the DOF transcription factor BPBF in the regulation of gibberellin-responsive genes in barley aleurone&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Mena&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 16%5BVolume%5D AND 53%5BPagination%5D AND Mena M%2C Vicente%2DCarbajosa J%2C Schmidt RJ%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Mena M, Vicente-Carbajosa J, Schmidt RJ, Carbonero P (1998) An endosperm-specific DOF protein from barley, highly conserved in wheat, binds to and activates transcription from the prolamin-box of a native B-hordein promoter in barley endosperm. Plant J 16: 53-62

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Mena&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=An endosperm-specific DOF protein from barley, highly conserved in wheat, binds to and activates transcription from the prolamin-box of a native B-hordein promoter in barley endosperm&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=An endosperm-specific DOF protein from barley, highly conserved in wheat, binds to and activates transcription from the prolamin-box of a native B-hordein promoter in barley endosperm&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Mena&as_ylo=1998&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 51%5BVolume%5D AND 352%5BPagination%5D AND Moreno%2DRisue�o MA%2C D�az I%2C Carrillo L%2C Fuentes R%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Moreno-Risue�o MA, D�az I, Carrillo L, Fuentes R, Carbonero P (2007) The HvDOF19 transcription factor mediates the abscisic acid-dependent repression of hydrolase genes in germinating barley aleurone. Plant J 51: 352-365

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Moreno-Risue�o&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The HvDOF19 transcription factor mediates the abscisic acid-dependent repression of hydrolase genes in germinating barley aleurone&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The HvDOF19 transcription factor mediates the abscisic acid-dependent repression of hydrolase genes in germinating barley aleurone&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Moreno-Risue�o&as_ylo=2007&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 53%5BVolume%5D AND 882%5BPagination%5D AND Moreno%2DRisue�o MA%2C Gonz�lez N%2C D�az I%2C Parcy F%2C Carbonero P%2C Vicente%2DCarbajosa J%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Moreno-Risue�o MA, Gonz�lez N, D�az I, Parcy F, Carbonero P, Vicente-Carbajosa J (2008) FUSCA3 from barley unveils a common transcriptional regulation of seed-specific genes between cereals and Arabidopsis. Plant J 53: 882-894

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Moreno-Risue�o&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=FUSCA3 from barley unveils a common transcriptional regulation of seed-specific genes between cereals and Arabidopsis&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=FUSCA3 from barley unveils a common transcriptional regulation of seed-specific genes between cereals and Arabidopsis&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Moreno-Risue�o&as_ylo=2008&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28J Exp Bot%5BTitle%5D%29 AND 52%5BVolume%5D AND 875%5BPagination%5D AND Nakamura S%2C Toyama T%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Nakamura S, Toyama T (2001) Isolation of a VP1 homologue from wheat and analysis of its expression in embryos of dormant and non-dormant cultivars. J Exp Bot 52: 875-876

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Nakamura&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Isolation of a VP1 homologue from wheat and analysis of its expression in embryos of dormant and non-dormant cultivars&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Isolation of a VP1 homologue from wheat and analysis of its expression in embryos of dormant and non-dormant cultivars&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Nakamura&as_ylo=2001&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Dev Biol%5BTitle%5D%29 AND 220%5BVolume%5D AND 412%5BPagination%5D AND Nambara E%2C Hayama R%2C Tsuchiya Y%2C Nishimura M%2C Kawaide H%2C Kamiya Y%2C Naito S%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Nambara E, Hayama R, Tsuchiya Y, Nishimura M, Kawaide H, Kamiya Y, Naito S (2000) The role of ABI3 and FUS3 loci in Arabidopsis thaliana on phase transition from late embryo development to germination. Dev Biol 220: 412-423

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Nambara&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The role of ABI3 and FUS3 loci in Arabidopsis thaliana on phase transition from late embryo development to germination&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The role of ABI3 and FUS3 loci in Arabidopsis thaliana on phase transition from late embryo development to germination&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Nambara&as_ylo=2000&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Mechanisms and genes involved in germination sensu stricto%2E%5BTitle%5D%29 AND Nonogaki H%2C Chen F%2C Bradford KJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Nonogaki H, Chen F, Bradford KJ (2007) Mechanisms and genes involved in germination sensu stricto. In Bradford K y Nonogaki H eds, Seed development, dormancy and germination. Blackwell Publishing, Oxford, pp 264-304

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Nonogaki&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Mechanisms and genes involved in germination sensu stricto.&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Mechanisms and genes involved in germination sensu stricto.&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Nonogaki&as_ylo=2007&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Barley%5BTitle%5D%29 AND 2%5BVolume%5D AND O�ate L%2C Vicente%2DCarbajosa J%2C Lara P%2C D�az I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=O�ate L, Vicente-Carbajosa J, Lara P, D�az I, Carbonero P (1999) Barley BLZ2, a seed-specific bZIP protein that interacts with BLZ1 in vivo and activates transcription from the GCN4-like motif of B-hordein promoters in barley endosperm. J Biol Chem 274: 9175-9182

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=O�ate&hl=en&c2coff=1


http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=O�ate&as_ylo=1999&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 6%5BVolume%5D AND 1567%5BPagination%5D AND Parcy F%2C Valon C%2C Raynal M%2C Gaubier%2DComella P%2C Delseny M%2C Giraudat J%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Parcy F, Valon C, Raynal M, Gaubier-Comella P, Delseny M, Giraudat J (1994) Regulation of gene expression programs during Arabidopsis seed development: roles of the ABI3 locus and of endogenous abscisic acid. Plant Cell 6: 1567-1582

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Parcy&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Regulation of gene expression programs during Arabidopsis seed development: roles of the ABI3 locus and of endogenous abscisic acid&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Regulation of gene expression programs during Arabidopsis seed development: roles of the ABI3 locus and of endogenous abscisic acid&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Parcy&as_ylo=1994&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28EMBO J%5BTitle%5D%29 AND 5%5BVolume%5D AND 829%5BPagination%5D AND Paz%2DAres J%2C Wienand U%2C Peterson PA%2C Saedler H%2E%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Paz-Ares J, Wienand U, Peterson PA, Saedler H. (1986) Molecular cloning of the c locus of Zea mays: a locus regulating the anthocyanin pathway. EMBO J 5: 829-833

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Paz-Ares&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular cloning of the c locus of Zea mays: a locus regulating the anthocyanin pathway&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular cloning of the c locus of Zea mays: a locus regulating the anthocyanin pathway&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Paz-Ares&as_ylo=1986&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 31%5BVolume%5D AND 639%5BPagination%5D AND Pritchard SL%2C Charlton WL%2C Baker A%2C Graham IA%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Pritchard SL, Charlton WL, Baker A, Graham IA (2002) Germination and storage reserve mobilization are regulated independently in Arabidopsis. Plant J. 31: 639-647

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Pritchard&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Germination and storage reserve mobilization are regulated independently in Arabidopsis&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Germination and storage reserve mobilization are regulated independently in Arabidopsis&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Pritchard&as_ylo=2002&as_allsubj=all&hl=en&c2coff=1


Rodríguez MV, Barrero JM, Corbineau F, Gubler F, Benech-Arnold RL (2015) Dormancy in cereals (not too much, not so little):about the mechanisms behind this trait. Seed Sci Res 25: 99-119


Rubio-Somoza I, Martinez M, Abraham Z, Diaz I, Carbonero P (2006a) Ternary complex formation between HvMYBS3 and otherfactors involved in transcriptional control in barley seeds. Plant J 47: 269-281


Rubio-Somoza I, Martinez M, Diaz I, Carbonero P (2006b) HvMCB1, a R1MYB transcription factor from barley with antagonisticregulatory functions during seed development and germination. Plant J 45: 17-30


Sun TP, Gubler F. (2004) Molecular mechanism of gibberellin signaling in plants. Annu Rev Plant Biol 55: 197-223.Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Suzuki M, Kao CY, Cocciolone S, McCarty DR (2001) Maize VP1 complements Arabidopsis abi3 and confers a novel ABA/auxininteraction in roots. Plant J 28:409-418.


Swaminathan K, Peterson K, Jack T (2008) The plant B3 superfamily. Trends Plant Sci 13: 647-655Pubmed: Author and TitleCrossRef: Author and TitleGoogle Scholar: Author Only Title Only Author and Title

Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. MolBiol Evol 30: 2725-2729


Thompson J, Higgins D, Gibson T (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignmentthrough sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22: 4673-4680


To A, Valon C, Savino G, Guilleminot J, Devic M, Giraudat J, Parcy F (2006) A network of local and redundant gene regulationgoverns Arabidopsis seed maturation. Plant Cell 18: 1642-1651


Vasil V, Marcotte WR Jr, Rosenkrans L, Cocciolone SM, Vasil IK, Quatrano RS, McCarty DR (1995) Overlap of Viviparous1 (VP1) andabscisic acid response elements in the Em promoter: G-box elements are sufficient but not necessary for VP1 transactivation.Plant Cell 7: 1511-1508


Vicente-Carbajosa J, Carbonero P (2005) Seed maturation: developing an intrusive phase to accomplish a quiescent state. Int JDev Biol 49: 645-651


Vicente-Carbajosa J, Moose SP, Parsons RL, Schmidt RJ (1997) A maize zinc-finger protein binds the prolamin box in zein genepromoters and interacts with the basic leucine zipper transcriptional activator Opaque2. Proc Natl Acad Sci USA 94: 7685-7690


Vicente-Carbajosa J, Oñate L, Lara P, Diaz I, Carbonero P (1998) Barley BLZ1: a bZIP transcriptional activator that interacts withendosperm-specific gene promoters. Plant J 13: 629-640


http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 21%5BVolume%5D AND 401%5BPagination%5D AND Reidt W%2C Wohlfarth T%2C Ellerstr�m M%2C Czihal A%2C Tewes A%2C Ezcurra I%2C Rask L%2C B�umlein H%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Reidt W, Wohlfarth T, Ellerstr�m M, Czihal A, Tewes A, Ezcurra I, Rask L, B�umlein H (2000) Gene regulation during late embryogenesis: the RY motif of maturation-specific gene promoters is a direct target of the FUS3 gene product. Plant J 21: 401-408

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Reidt&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Gene regulation during late embryogenesis: the RY motif of maturation-specific gene promoters is a direct target of the FUS3 gene product&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Gene regulation during late embryogenesis: the RY motif of maturation-specific gene promoters is a direct target of the FUS3 gene product&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Reidt&as_ylo=2000&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Seed Sci Res%5BTitle%5D%29 AND 25%5BVolume%5D AND 99%5BPagination%5D AND Rodr�guez MV%2C Barrero JM%2C Corbineau F%2C Gubler F%2C Benech%2DArnold RL%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Rodr�guez MV, Barrero JM, Corbineau F, Gubler F, Benech-Arnold RL (2015) Dormancy in cereals (not too much, not so little): about the mechanisms behind this trait. Seed Sci Res 25: 99-119

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Rodr�guez&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Dormancy in cereals (not too much, not so little): about the mechanisms behind this trait&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Dormancy in cereals (not too much, not so little): about the mechanisms behind this trait&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Rodr�guez&as_ylo=2015&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 47%5BVolume%5D AND 269%5BPagination%5D AND Rubio%2DSomoza I%2C Martinez M%2C Abraham Z%2C Diaz I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Rubio-Somoza I, Martinez M, Abraham Z, Diaz I, Carbonero P (2006a) Ternary complex formation between HvMYBS3 and other factors involved in transcriptional control in barley seeds. Plant J 47: 269-281

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Rubio-Somoza&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Ternary complex formation between HvMYBS3 and other factors involved in transcriptional control in barley seeds&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Ternary complex formation between HvMYBS3 and other factors involved in transcriptional control in barley seeds&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Rubio-Somoza&as_ylo=2006&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28HvMCB%5BTitle%5D%29 AND 1%5BVolume%5D AND a R1MYB transcription factor from barley with antagonistic regulatory functions during seed development and germination%2E Plant J 45%3A 17%5BPagination%5D AND Rubio%2DSomoza I%2C Martinez M%2C Diaz I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Rubio-Somoza I, Martinez M, Diaz I, Carbonero P (2006b) HvMCB1, a R1MYB transcription factor from barley with antagonistic regulatory functions during seed development and germination. Plant J 45: 17-30

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Rubio-Somoza&hl=en&c2coff=1


http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Rubio-Somoza&as_ylo=2006&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Annu Rev Plant Biol%5BTitle%5D%29 AND 55%5BVolume%5D AND 197%5BPagination%5D AND Sun TP%2C Gubler F%2E%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Sun TP, Gubler F. (2004) Molecular mechanism of gibberellin signaling in plants. Annu Rev Plant Biol 55: 197-223.

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Sun&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular mechanism of gibberellin signaling in plants&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Molecular mechanism of gibberellin signaling in plants&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Sun&as_ylo=2004&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant J%5BTitle%5D%29 AND 28%5BVolume%5D AND 409%5BPagination%5D AND Suzuki M%2C Kao CY%2C Cocciolone S%2C McCarty DR%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Suzuki M, Kao CY, Cocciolone S, McCarty DR (2001) Maize VP1 complements Arabidopsis abi3 and confers a novel ABA/auxin interaction in roots. Plant J 28:409-418.

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Suzuki&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Maize VP1 complements Arabidopsis abi3 and confers a novel ABA/auxin interaction in roots&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Maize VP1 complements Arabidopsis abi3 and confers a novel ABA/auxin interaction in roots&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Suzuki&as_ylo=2001&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Trends Plant Sci%5BTitle%5D%29 AND 13%5BVolume%5D AND 647%5BPagination%5D AND Swaminathan K%2C Peterson K%2C Jack T%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Swaminathan K, Peterson K, Jack T (2008) The plant B3 superfamily. Trends Plant Sci 13: 647-655

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Swaminathan&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The plant B3 superfamily&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The plant B3 superfamily&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Swaminathan&as_ylo=2008&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28MEGA%5BTitle%5D%29 AND 6%5BVolume%5D AND Molecular Evolutionary Genetics Analysis version 6%2E0%2E Mol Biol Evol 30%3A 2725%5BPagination%5D AND Tamura K%2C Stecher G%2C Peterson D%2C Filipski A%2C Kumar S%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol Biol Evol 30: 2725-2729

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Tamura&hl=en&c2coff=1


http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Tamura&as_ylo=2013&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Nucleic Acids Res%5BTitle%5D%29 AND 22%5BVolume%5D AND 4673%5BPagination%5D AND Thompson J%2C Higgins D%2C Gibson T%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Thompson J, Higgins D, Gibson T (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22: 4673-4680

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Thompson&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Thompson&as_ylo=1994&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 18%5BVolume%5D AND 1642%5BPagination%5D AND To A%2C Valon C%2C Savino G%2C Guilleminot J%2C Devic M%2C Giraudat J%2C Parcy F%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=To A, Valon C, Savino G, Guilleminot J, Devic M, Giraudat J, Parcy F (2006) A network of local and redundant gene regulation governs Arabidopsis seed maturation. Plant Cell 18: 1642-1651

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=To&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=A network of local and redundant gene regulation governs Arabidopsis seed maturation&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A network of local and redundant gene regulation governs Arabidopsis seed maturation&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=To&as_ylo=2006&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell%5BTitle%5D%29 AND 7%5BVolume%5D AND 1511%5BPagination%5D AND Vasil V%2C Marcotte WR Jr%2C Rosenkrans L%2C Cocciolone SM%2C Vasil IK%2C Quatrano RS%2C McCarty DR%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Vasil V, Marcotte WR Jr, Rosenkrans L, Cocciolone SM, Vasil IK, Quatrano RS, McCarty DR (1995) Overlap of Viviparous1 (VP1) and abscisic acid response elements in the Em promoter: G-box elements are sufficient but not necessary for VP1 transactivation. Plant Cell 7: 1511-1508

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Vasil&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Overlap of Viviparous1 (VP1) and abscisic acid response elements in the Em promoter: G-box elements are sufficient but not necessary for VP1 transactivation&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Overlap of Viviparous1 (VP1) and abscisic acid response elements in the Em promoter: G-box elements are sufficient but not necessary for VP1 transactivation&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Vasil&as_ylo=1995&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Int J Dev Biol%5BTitle%5D%29 AND 49%5BVolume%5D AND 645%5BPagination%5D AND Vicente%2DCarbajosa J%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Vicente-Carbajosa J, Carbonero P (2005) Seed maturation: developing an intrusive phase to accomplish a quiescent state. Int J Dev Biol 49: 645-651

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Vicente-Carbajosa&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Seed maturation: developing an intrusive phase to accomplish a quiescent state&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Seed maturation: developing an intrusive phase to accomplish a quiescent state&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Vicente-Carbajosa&as_ylo=2005&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Proc Natl Acad Sci USA%5BTitle%5D%29 AND 94%5BVolume%5D AND 7685%5BPagination%5D AND Vicente%2DCarbajosa J%2C Moose SP%2C Parsons RL%2C Schmidt RJ%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Vicente-Carbajosa J, Moose SP, Parsons RL, Schmidt RJ (1997) A maize zinc-finger protein binds the prolamin box in zein gene promoters and interacts with the basic leucine zipper transcriptional activator Opaque2. Proc Natl Acad Sci USA 94: 7685-7690


http://scholar.google.com/scholar?as_q=A maize zinc-finger protein binds the prolamin box in zein gene promoters and interacts with the basic leucine zipper transcriptional activator Opaque2&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A maize zinc-finger protein binds the prolamin box in zein gene promoters and interacts with the basic leucine zipper transcriptional activator Opaque2&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Vicente-Carbajosa&as_ylo=1997&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Barley%5BTitle%5D%29 AND 1%5BVolume%5D AND Vicente%2DCarbajosa J%2C O�ate L%2C Lara P%2C Diaz I%2C Carbonero P%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Vicente-Carbajosa J, O�ate L, Lara P, Diaz I, Carbonero P (1998) Barley BLZ1: a bZIP transcriptional activator that interacts with endosperm-specific gene promoters. Plant J 13: 629-640



http://scholar.google.com/scholar?as_q=&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Vicente-Carbajosa&as_ylo=1998&as_allsubj=all&hl=en&c2coff=1

Yan D, Duermeyer L, Leoveanu C, Nambar E (2014) The functions of the endosperm during seed germination. Plant Cell Physiol55: 1521-1533


Zeng Y, Raimondi N, Kermode AR (2003) Role of an ABI3 homologue in dormancy maintenance of yellow-cedar seeds and in theactivation of storage protein and Em gene promoters. Plant Mol Biol 51: 39-49


Zeng Y, Zhao T, Kermode AR (2013) A conifer ABI3-interacting protein plays important roles during key transitions of the plant lifecycle. Plant Physiol 161: 179-195


http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Cell Physiol%5BTitle%5D%29 AND 55%5BVolume%5D AND 1521%5BPagination%5D AND Yan D%2C Duermeyer L%2C Leoveanu C%2C Nambar E%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Yan D, Duermeyer L, Leoveanu C, Nambar E (2014) The functions of the endosperm during seed germination. Plant Cell Physiol 55: 1521-1533

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Yan&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=The functions of the endosperm during seed germination&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=The functions of the endosperm during seed germination&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Yan&as_ylo=2014&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Mol Biol%5BTitle%5D%29 AND 51%5BVolume%5D AND 39%5BPagination%5D AND Zeng Y%2C Raimondi N%2C Kermode AR%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Zeng Y, Raimondi N, Kermode AR (2003) Role of an ABI3 homologue in dormancy maintenance of yellow-cedar seeds and in the activation of storage protein and Em gene promoters. Plant Mol Biol 51: 39-49

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Zeng&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=Role of an ABI3 homologue in dormancy maintenance of yellow-cedar seeds and in the activation of storage protein and Em gene promoters&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=Role of an ABI3 homologue in dormancy maintenance of yellow-cedar seeds and in the activation of storage protein and Em gene promoters&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Zeng&as_ylo=2003&as_allsubj=all&hl=en&c2coff=1

http://www.ncbi.nlm.nih.gov/pubmed?term=%28Plant Physiol%5BTitle%5D%29 AND 161%5BVolume%5D AND 179%5BPagination%5D AND Zeng Y%2C Zhao T%2C Kermode AR%5BAuthor%5D&dopt=abstract

http://search.crossref.org/?page=1&rows=2&q=Zeng Y, Zhao T, Kermode AR (2013) A conifer ABI3-interacting protein plays important roles during key transitions of the plant life cycle. Plant Physiol 161: 179-195

http://scholar.google.com/scholar?as_q=&num=10&as_occt=any&as_sauthors=Zeng&hl=en&c2coff=1

http://scholar.google.com/scholar?as_q=A conifer ABI3-interacting protein plays important roles during key transitions of the plant life cycle&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=any&as_publication=&as_yhi=&as_allsubj=all&hl=en&lr=&c2coff=1

http://scholar.google.com/scholar?as_q=A conifer ABI3-interacting protein plays important roles during key transitions of the plant life cycle&num=10&btnG=Search+Scholar&as_occt=any&as_sauthors=Zeng&as_ylo=2013&as_allsubj=all&hl=en&c2coff=1

Documents

A developmental switch of gene expression in the barley seed …€¦ · 26/02/2016 · The deduced HvVP1 protein sequence from cv Igri is more than 99% 161 identical with those