JGI: Genome size impacts on plant adaptation

Adaptation in plant genomes: a role for genome size?

Jeffrey Ross-Ibarra @jrossibarra • www.rilab.org

Dept. Plant Sciences • Center for Population Biology • Genome Center University of California Davis

photo by lady_lbrty

Kew C-Value Database

Gaut and Ross-Ibarra 2008

Paris Japonica150GB Genome

Genlisia aurea63MB Genome Michal Rubeš

wide variation in genome size in plants


what explains genome size variation?

Lynch & Connery 2003 ScienceWhitney et al. 2010 Evolution


what explains genome size variation?

Lynch & Connery 2003 ScienceWhitney et al. 2010 Evolution

seed weight

Knight et al 2005 AoB

genome size

geno

me

size

leaf

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acorrelates of genome size: phenotypes across species

elevation

geno

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Bilinski et al. In Prep

correlates of genome size: altitude within Zea mays

0

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genome size

late floweringearly flowering

Rayburn et al. 1994 Plant Breeding

# pl

ants

hard sweep

how do genomes adapt?

hard sweep


hard sweep


hard sweep

multiple mutations

standing variation

“soft” sweeps


hard sweep

multiple mutations

polygenic adaptation

standing variation

“soft” sweeps


M T G P H R L

GGTCGAC ATG ACT GGT CCA CAT CGA CTG TAG

M T G P H R L

GGTCGAC ATG ACT GGT CCA CAT CGA CTG TAG

M T N P H R L

GGTCGAC ATG ACT GAT CCA CAT CGA CTG TAG

structural change to protein

M T G P H R L

GGTAAAC ATG ACT GGT CCA CAT CGA CTG TAG

GG—-AC ATG ACT GGT CCA CAT CGA CTG TAG

regulatory change to expression

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Figure 1 _ Main Text

Tenaillon et al. 2010 TIP Springer et al. 2016 Plant Cell

Tenaillon et al. 2010 TIP Springer et al. 2016 Plant Cell

Ne individuals, µ beneficial mutation rate per trait

bigger genome, larger mutation target, higher µ

selection from standing variation when 2Neµ > 1

predict that larger genomes adapt via noncoding changes, standing variation

Hancock et al 2011 Science

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esArabidopsis adaptation predominantly coding

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maize (2.5Gb)Arabidopsis

log 1C genome size

Suketoshi

how does adaptation work in maize?

maizeteosinte

standing variation

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NATURE GENETICS ADVANCE ONLINE PUBLICATION 3

L E T T E R S

mutation rate21, strongly suggesting that the Hopscotch insertion (and thus, the older Tourist as well) existed as standing genetic variation in the teosinte ancestor of maize. Thus, we conclude that the Hopscotch insertion likely predated domestication by more than 10,000 years and the Tourist insertion by an even greater amount of time.

We identified four fixed differences in the portion of the proximal and distal components of the control region that show evidence of selection. We used transient assays in maize leaf protoplasts to test all four differences for effects on gene expression. Maize and teosinte chromosomal segments for the portions of the proximal and distal components with these four differences were cloned into reporter constructs upstream of the minimal promoter of the cauliflower mosaic virus (mpCaMV), the firefly luciferase ORF and the nopaline synthase (NOS) terminator (Fig. 4). Each construct was assayed for luminescence after transformation by electroporation into maize pro-toplast. The constructs for the distal component contrast the effects of the Tourist insertion plus the single fixed nucleotide substitution that distinguish maize and teosinte. Both the maize and teosinte constructs for the distal component repressed luciferase expression

relative to the minimal promoter alone. The maize construct with Tourist excised gave luciferase expression equivalent to the native maize and teosinte constructs and less expression than the minimal promoter alone. These results indicate that this segment is function-ally important, acting as a repressor of luciferase expression and, by inference, of tb1 expression in vivo. However, we did not observe any difference between the maize and teosinte constructs as anticipated. One possible cause for the lack of differences in expression between the maize and teosinte constructs might be that additional proteins required to cause these differences are not present in maize leaf pro-toplast. Another possibility is that the factor affecting phenotype in the distal component lies in the unselected region between −64.8 and −69.5 kb, which is not included in the construct. Nevertheless, the results do indicate that the distal component has a functional element that acts as a repressor. The functional importance of this segment is supported by its low level of nucleotide diversity (Fig. 3a), suggesting a history of purifying selection.

The constructs for the proximal component of the control region contrast the effects of the Hopscotch insertion plus a single fixed nucleo-tide substitution that distinguish maize and teosinte. The construct with the maize sequence including Hopscotch increased expression of the luciferase reporter twofold relative to the teosinte construct for the proximal control region and the minimal promoter alone (Fig. 4). Luciferase expression was returned to the level of the teosinte con-struct and the minimal promoter construct by deleting the Hopscotch element from the full maize construct. These results indicate that the Hopscotch element enhances luciferase expression and, by

a

b

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A B C D M

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P = 0.95 P = 0.41 P = 0.04

HKA neutrality tests

P 0.0001

0.04

0.02

0–67 kb –66 kb

Distalcomponent

Teosinte clusterhaplotype

Maize clusterhaplotype

Proximalcomponent

–65 kbTourist408 bp

Hopscotch4,885 bp

–64 kb –58 kb

Figure 3 Sequence diversity in maize and teosinte across the control region. (a) Nucleotide diversity across the tb1 upstream control region. Base-pair positions are relative to AGPv2 position 265,745,977 of the maize reference genome sequence. P values correspond to HKA neutrality tests for regions A–D, as defined by the dotted lines. Green shading signifies evidence of neutrality, and pink shading signifies regions of non-neutral evolution. Nucleotide diversity ( ) for maize (yellow line) and teosinte (green line) were calculated using a 500-bp sliding window with a 25-bp step. The distal and proximal components of the control region with four fixed sequence differences between the most common maize haplotype and teosinte haplotype are shown below. (b) A minimum spanning tree for the control region with 16 diverse maize and 17 diverse teosinte sequences. Size of the circles for each haplotype group (yellow, maize; green, teosinte) is proportional to the number of individuals within that haplotype.

Transient assay constructs

mpCaMV luc

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luc

luc

luc

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luc

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Tourist

mpCaMV

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Figure 4 Constructs and corresponding normalized luciferase expression levels. Transient assays were performed in maize leaf protoplast. Each construct is drawn to scale. The construct backbone consists of the minimal promoter from the cauliflower mosaic virus (mpCaMV, gray box), luciferase ORF (luc, white box) and the nopaline synthase terminator (black box). Portions of the proximal and distal components of the control region (hatched boxes) from maize and teosinte were cloned into restriction sites upstream of the minimal promoter. “ ” denotes the excision of either the Tourist or Hopscotch element from the maize construct. Horizontal green bars show the normalized mean with s.e.m. for each construct.

relative expressionconstructStuder et al. 2011 Nat. Gen.; Vann et al. 2015 PeerJ

enhances expression

teosinte branched - tb1

hard sweep

Figure 1.Phenotypes. a. Maize ear showing the cob (cb) exposed at top. b. Teosinte ear with the rachisinternode (in) and glume (gl) labeled. c. Teosinte ear from a plant with a maize allele of tga1introgressed into it. d. Close-up of a single teosinte fruitcase. e. Close-up of a fruitcase fromteosinte plant with a maize allele of tga1 introgressed into it. f. Ear of maize inbred W22(Tga1-maize allele) with the cob exposed showing the small white glumes at the base. g. Earof maize inbred W22:tga1 which carries the teosinte allele, showing enlarged (white) glumes.h. Ear of maize inbred W22 carrying the tga1-ems1 allele, showing enlarged glumes. For highermagnification copies of f–h see Supplementary Information.

Wang et al. Page 10

Nature. Author manuscript; available in PMC 2006 May 23.

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anuscriptWang et al. 2015 Genetics

protein change

teosinte glume architecture - tga1

multiple mutations

Wills et al. 2013 PLoS Genetics

teosinte maizeClint Whipple, BYU

grassy tillers - gt1

5’ control region 3’ UTRmodifies expression

hard sweep

M T N P H R L

GGTCGA ATG ACT GAT CCA CAT CGA CTG TAG

tga1 gt1 tb1

Multiple Mutations

Standing Variation

M T G P H R L

GGTAAA ATG ACT GGT CCA CAT CGA CTG TAG

Hufford et al. 2012 Nat. Gen. Chia et al. 2012 Nat. Gen

13 teosinte 23 maizegenomes:

genome-wide evidence of adaptation






5-10% selected regions do not include genes


whereas others are lost after domestication (Fig. 3B). It should benoted that many of these genes have unique coexpression edges inmaize that are not observed in teosinte (Fig. S4B).

Expression data provide an opportunity to investigate furtherfunctional alterations to genes located within genomic regionsthat population genomic analyses identify as targets of selective

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DE(n=612)

AEC(n=1115)

Dom/Imp genes(n=1761)

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Teosinte network edges Maize network edges

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GRMZM2G068436

GRMZM2G137947

GRMZM2G375302

Mb

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Fig. 3. Analysis of genes with altered expression or conservation and targets of selection during improvement and/or domestication. (A) Venn diagramshowing the overlap between DE genes, AEC genes, and the genes that occur in genomic regions that have evidence for selective sweeps during maizedomestication or improvement (Dom/Imp genes). (B) Teosinte coexpression networks for three genes (GRMZM2G068436, GRMZM2G137947, andGRMZM2G375302). (Right) Edges that are maintained in maize coexpression networks are shown. Although the differentially expressed gene (red node) ishighly connected in teosinte, most of these connections are lost in maize. However, some parts of the teosinte network are still conserved in maize. (C) Cross-population composite likelihood ratio test (XP-CLR) plot shows the evidence for a selective sweep that occurs on chromosome 9. The tick marks along the xaxis represent genes, and the red tick mark indicates the gene (GRMZM2G448355) that was chosen as the candidate target of selection and is differentiallyexpressed in maize and teosinte. The bar plot underneath the graph shows the expression levels of all maize (blue) and teosinte (red) samples. (D) XP-CLR plotfor a large region on chromosome 5. The candidate target of selection is indicated in green and shows similar expression in maize and teosinte. Two othergenes (red) exhibit DE. (E) Neighbor-joining tree shows the relationships among the haplotypes at GRMZM2G141858. (Right) Bar plot shows expression levelsfor each genotype; red bars indicate teosinte genotypes, and blue bars represent maize genotypes. At least one teosinte genotype (TIL15) contains thehaplotype that has been selected in maize and has expression levels similar to maize genotypes.

Table 2. Genes in selected regions with evidence for DE or AEC

Gene listNo. genes selectedduring dom/imp

% up-regulatedin maize Significance

% higher connectedin maize % candidates

AEC and DE (n = 276) 46 76 0.0002 41.3 39.1DE only (n = 336) 44 61 0.0230 40.9 22.7AEC only (n = 839) 89 54 0.1837 57.3 32.6

dom, domestication; imp, improvement.

4 of 6 | www.pnas.org/cgi/doi/10.1073/pnas.1201961109 Swanson-Wagner et al.

ExpressionGenealogy

teosintemaize

• ~500 selected regions

• 11M shared vs 3000 fixed SNPs

• show differential expression, decreased expression variation

selection on regulatory sequence, standing variation

Hufford et al. 2012 Nat. Gen. Swanson-Wagner et al. 2012 PNAS

Beissinger et al. BioRxiv

nucl

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distance to nearest substitution (cM)

hard sweeps in genes play minor role in maize


nucl

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distance to nearest substitution (cM)

hard sweeps in genes play minor role in maize

Wallace et al. 2014 PLoS Genetics

QTL alleles enriched for noncoding

Rodgers-Melnick et al. 2016 PNAS

Variance PartitioningGWAS candidate SNPs

Makarevitch et al. 2015 PLoS Genetics


single TE family many genes


single TE family many genes

new insertions activate expression

Makarevitch et al. 2014 bioRxiv

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Lines without the TE insertion

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single gene, many individuals

how to adapt: Zea mays

M T G P H R L

GGTAAA ATG ACT GGT CCA CAT CGA CTG TAG

regulatory variation (including TEs)multiple

mutations

“soft” sweeps

standing variation

Sattah et al. 2011 PLoS Gen. Williamson et al. 2014 PLoS Gen Hernandez et al. 2011 ScienceRoss-Ibarra et al. 2009 Genetics

Sattah et al. 2011 PLoS Gen. Williamson et al. 2014 PLoS Gen Hernandez et al. 2011 ScienceRoss-Ibarra et al. 2009 Genetics

Sattah et al. 2011 PLoS Gen. Williamson et al. 2014 PLoS Gen Hernandez et al. 2011 Science

dive

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distance from substitution

Ross-Ibarra et al. 2009 Genetics


dive

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distance from substitution

20% nonsyn. adaptive 10% nonsyn. adaptive

50% nonsyn. adaptive 40% nonsyn. adaptive

Ross-Ibarra et al. 2009 Genetics

Ne effective number of diploid individuals

s selection coefficient

selection is effective if 2Nes > 1

differences in adaptation due to drift and small population size?

0.05Na

Na

Na3NaNe ~ 450,000


0.05Na

Na

Na3NaNe ~ 450,000


Ne ~ 1,000,000

0.05Na

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Na3NaNe ~ 450,000


Ne ~ 1,000,000

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popBKN_4HapBKN_6HapTIL_4Hap_JaliscoTIL_6Hap

Ne ~ 1,000,000,000

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Ne ~ 1,000,000

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popBKN_4HapBKN_6HapTIL_4Hap_JaliscoTIL_6Hap

Ne ~ 1,000,000,000

Ne ~ 5,000,000,000


dive

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Ne >> 1,000,000 Ne ~ 10,000*

Ne ~ 2,000,000 Ne ~ 600,000


dive

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µ ∝ 2,500 Mbp µ ∝ 3,100 Mbp

µ ∝ 130 Mbp µ ∝ 220 Mbp

Pyhäjärvi et al. GBE 2013

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large genomes enriched in noncoding adaptive variants

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Hancock et al 2011 Science Fraser et al. 2013 Gen. Research

Pyhäjärvi et al. GBE 2013

large genomes enriched in noncoding adaptive variants

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Hancock et al 2011 Science Fraser et al. 2013 Gen. Research

• Adaptation in maize occurs from standing variation and targets regulatory variants

• Large genomes may have more targets, more standing variation, and more regulatory adaptation

• Efforts to identify functional variation should consider genome size in designing experiments and genotyping

Genome Size and Adaptation


Acknowledgments

Maize Diversity GroupPeter Bradbury

Ed Buckler John Doebley Theresa Fulton

Sherry Flint-Garcia Jim Holland

Sharon Mitchell Qi Sun

Doreen Ware

CollaboratorsCSI Davis

Nathan Springer

Lab AlumniTim Beissinger (USDA-ARS, Mizzou)

Kate Crosby (Monsanto) Matt Hufford (Iowa State)

Tanja Pyhäjärvi (Oulu) Shohei Takuno (Sokendai)

Joost van Heerwaarden (Wageningen)