Decoding options and accuracy of translation of developmentally regulated UUA codon in Streptomyces: bioinformatic analysisIhor Rokytskyy†, Oksana Koshla†, Victor Fedorenko and Bohdan Ostash*

FindingsThe bldA mutants of Streptomyces coelicolor A3(2) were first isolated almost 40 years ago (Merrick 1976) and 11 years later were shown to carry mutations within gene for leucyl tRNA

LeuUAA. (Lawlor et  al. 1987) This mutation abolishes aerial mycelium formation

(Bald phenotype) and antibiotic production by streptomycetes; currently bldA is exten-sively used as a tool to activate cryptic secondary metabolome (Hackl and Bechthold

Abstract

Background: The gene bldA for leucyl tRNALeuUAA is known for almost 30 years as a key regulator of morphogenesis and secondary metabolism in genus Streptomy-ces. Codon UUA is the rarest one in Streptomyces genomes and is present exclusively in genes with auxiliary functions. Delayed accumulation of translation-competent tRNA

LeuUAA is believed to confine the expression of UUA-containing transcripts to

stationary phase. Implicit to the regulatory function of UUA codon is the assumption about high accuracy of its translation, e.g. the latter should not occur in the absence of cognate tRNALeuUAA. However, a growing body of facts points to the possibility of mistranslation of UUA-containing transcripts in the bldA-deficient mutants. It is not known what type of near-cognate tRNA(s) may decode UUA in the absence of cognate tRNA in Streptomyces, and whether UUA possesses certain inherent properties (such as increased/decreased accuracy of decoding) that would favor its use for regulatory purposes.

Findings: Here we took bioinformatic approach to address these questions. We cata-logued the entire complement of tRNA genes from several relevant Streptomyces and identified genes for posttranscriptional modifications of tRNA that might be involved in UUA decoding by cognate and near-cognate tRNAs.

Conclusions: Based on tRNA gene content in Streptomyces genomes, we propose possible scenarios of UUA codon mistranslation. UUA is not associated with an increased rate of missense errors as compared to other leucyl codons, contrasting gen-eral belief that low-abundant codons are more error-prone than the high-abundant ones.

Keywords: Streptomyces, bldA, tRNA, UUA codon, Codons, Models of translation

Open Access

© 2016 The Author(s). This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

SHORT REPORT

Rokytskyy et al. SpringerPlus (2016) 5:982 DOI 10.1186/s40064-016-2683-6

*Correspondence: b.ostash@lnu.edu.ua †Ihor Rokytskyy and Oksana Koshla have contributed equally to this work

Department of Genetics and Biotechnology, Ivan Franko National University of Lviv, Hrushevskoho st. 4, Lviv 79005, Ukraine

Page 2 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

2015). Codon TTA, whose decoding is controlled by bldA, is very rare in GC-rich Strep-tomyces genomes, and present only in genes with unknown and auxiliary functions, such as colony morphological development and antibiotic production. As accumulation of translation-competent, charged tRNA

LeuUAA is confined to late stages of growth, so

does the expression of TTA-containing genes (Chater 2006). UUA codon and its cognate tRNA were long time ago suggested to form a genetic switch that operates at the level of translation (Hopwood 1987). Use of TTA for regulatory purposes is somewhat contro-versial. On one hand, rarity of this codon ensures that only certain genes are influenced by bldA-based switch. On the other hand, rare codons are thought to be associated with higher missense error rates, which would not favor their proper operation as a switch. Although it was suggested implicitly that UUA is decoded accurately, there is a number of notable exceptions. Particularly, bldA mutants showed no Bald phenotype on certain solid media (Hopwood 1987); several TTA-containing genes were expressed in bldA-deficient strains (Trepanier et  al. 2002; Makitrynskyy et  al. 2013), particularly when their transcription is artificially elevated (Gramajo et  al. 1993). All these observations imply that efficient mistranslation of UUA codon is possible at least under some condi-tions. It is not known what tRNAs could potentially recognize UUA in the absence of tRNA

LeuUAA and what structural and functional peculiarities of tRNA

LeuUAA contribute

to its regulatory function (Pettersson and Kirsebom 2011). Here we took bioinformatic approach to obtain new insight into this issue and to chart new directions for experi-mental verification.

Although there are several databases of tRNA genes, such as GtRNAdb and tRNADB-CE, they provide contradictory information on tRNA content for model strain Strep-tomyces coelicolor A3(2) and lack data on several species relevant to this work. Furthermore, available online resources do not show what kind of tRNA may decode certain codon via wobble interaction. We therefore compiled all available information on tRNA genes and their decoding capacity for six Streptomyces species with known cases of bldA-based regulation using several databases and search tools detailed in Additional file 1. It could be concluded that overall tRNA gene content is highly conserved in six analyzed Streptomyces genomes, although copy number of individual tRNA genes var-ies (Table  1). For several codons there were no acceptor tRNAs (for example, alanine codon GCT); those apparently are recognized by isoacceptor tRNAs (e.g. GCT is read by GCC isoacceptor; see Table 1), which are encoded within the analyzed genomes. It is common for all known organisms that the entire set of sense codons (61 + 1 initiator) is read by far fewer than 62 isoacceptors; extreme cases of anticodon-sparing are docu-mented in some archaea and mycoplasmas, where only 26–33 anticodons are required to read the genetic code (Marck and Grosjean 2002). All genomes contain single tRNA gene for UUA decoding. Therefore, differences in bldA mutant phenotypes across dif-ferent species could not be ascribed to variations in tRNA gene content. Codon UUA could be poorly recognized by phenylalanine tRNA

PheAAA via wobble interactions (Lim

and Curran 2001). However, no respective tRNA gene is present in all studied Strepto-myces genomes. Cytidine posttranscriptionally modified with lysidine (k2C34) is known to recognize adenosine in third codon position. This kind of modification to date was described only for anticodon CAU, which normally decodes methionine codon AUG. The k2C34-containing tRNACAU is charged with isoleucine and recognizes isoleucine

Page 3 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Tabl

e 1

tRN

A g

enes

in s

ix S

trep

tom

yces

gen

omes

AA

Codo

nA

ntic

odon

Codo

n-

antic

odon

re

cogn

ition

tRN

A g

enes

*

A3(

2)63

**J1

074

65AT

CC 1

0712

65TK

2461

ATCC

146

7268

ATCC

270

6466

12

34

56

78

910

Ala

(A)

GC

TAG

CA

GC

, GG

C–

––

––

–

GCC

GG

CA

GC

, GG

CSC

Ot1

4, S

COt1

5XN

R_43

62, X

NR_

4366

SVEN

_t15

, SVE

N_t

16,

SVEN

_t56

SLI_

1001

4, S

LI_1

0015

SSFG

_RS2

3155

, SS

FG_R

S231

65SC

N_R

S083

55,

SCN

_RS0

8365

GC

ATG

CA

GC

, TG

CSC

Ot3

2XN

R_30

21SV

EN_t

34SL

I_10

033

SSFG

_RS1

7340

SCN

_RS1

4115

GCG

CGC

TGC

, CG

CSC

Ot2

6XN

R_21

23SV

EN_t

28SL

I_10

026

SSFG

_RS3

3025

, SS

FG_R

S202

10SC

N_R

S111

30

Val (

V)

GTT

AAC

AA

C, G

AC

––

––

––

GTC

GAC

AA

C, G

AC

vala

, val

b, v

alg

XNR_

5303

, XN

R_53

04,

XNR_

5305

SVEN

_t6,

SVE

N_t

7,

SVEN

_t8

SLI_

1000

7SS

FG_R

S282

15,

SSFG

_RS2

8210

, SS

FG_R

S282

20

SCN

_RS0

3735

, SC

N_R

S037

25,

SCN

_RS0

3730

, SC

N_R

S045

30

GTA

TAC

AA

C, T

AC

SCO

t10

XNR_

4542

SVEN

_t11

SLI_

1001

0SS

FG_R

S242

05SC

N_R

S076

25

GTG

CAC

TAC

, CA

CSC

Ot0

1, S

COt0

2XN

R_53

17, X

NR_

5319

SVEN

_t2,

SVE

N_t

3SL

I_10

003,

SLI

_100

04SS

FG_R

S283

10,

SSFG

_RS2

8275

,SC

N_R

S036

55,

SCN

_RS0

3665

Thr (

T)A

CT

AGT

AG

T, G

GT

––

––

––

AC

ATG

TA

GT,

TG

TSC

Ot4

7XN

R_25

51SV

EN_t

49SL

I_10

051

SSFG

_RS1

9050

SCN

_RS1

5670

ACC

GG

TA

GT,

GG

TSC

Ot4

9, S

COt6

1XN

R_11

17, X

NR_

3695

SVEN

_t53

, SVE

N_t

64SL

I_10

053,

SLI

_100

65SS

FG_R

S143

20SC

N_R

S175

50,

SCN

_RS2

2525

ACG

CGT

TGT,

CG

TSC

Ot2

8XN

R_26

97SV

EN_t

30SL

I_10

029

SSFG

_RS1

5630

SCN

_RS1

1750

Pro

(P)

CCT

AGG

AG

G, G

GG

––

––

––

CCC

GG

GA

GG

, GG

GSC

Ot6

3, S

COt6

4,

SCO

t65

XNR_

0211

, XN

R_02

44SV

EN_t

67, S

VEN

_t68

SLI_

1006

7, S

LI_1

0068

, SL

I_10

069

SSFG

_RS0

3965

, SS

FG_R

S040

60,

SSFG

_RS5

0385

0

SCN

_RS2

5645

, SC

N_R

S261

15

CCA

TGG

AG

G, T

GG

SCO

t16

XNR_

4324

SVEN

_t17

SLI_

1001

6SS

FG_R

S229

15SC

N_R

S085

55

CCG

CGG

TGG

, CG

GSC

Ot2

9XN

R_27

39SV

EN_t

31SL

I_10

030

SSFG

_RS1

5825

SCN

_RS1

2385

Page 4 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Tabl

e 1

cont

inue

d

AA

Codo

nA

ntic

odon

Codo

n-

antic

odon

re

cogn

ition

tRN

A g

enes

*

A3(

2)63

**J1

074

65AT

CC 1

0712

65TK

2461

ATCC

146

7268

ATCC

270

6466

12

34

56

78

910

Ser (

S)TC

TAG

AA

GA

, GG

A–

––

––

–

TCC

GG

AA

GA

, GG

ASC

Ot3

8XN

R_28

59SV

EN_t

40SL

I_10

039

SSFG

_RS1

8130

SCN

_RS1

4915

TCA

TGA

AG

A, T

GA

SCO

t34

XNR_

2906

SVEN

_t36

SLI_

1003

5SS

FG_R

S178

45SC

N_R

S136

55

TCG

CGA

TGA

, CG

ASC

Ot3

7XN

R_28

82SV

EN_t

39SL

I_10

038

SSFG

_RS1

8035

SCN

_RS1

4685

Ser (

Z)A

GT

ACT

AC

T, G

CT

––

––

––

AG

CG

CTG

CT

SCO

t35

XNR_

2899

SVEN

_t37

SLI_

1003

6SS

FG_R

S179

10SC

N_R

S135

80

Phe

(F)

TTT

AAA

AA

A, G

AA

––

––

––

TTC

GAA

GA

ASC

Ot4

2XN

R_28

33SV

EN_t

44SL

I_10

043

SSFG

_RS1

8285

SCN

_RS1

5085

Met

(M)

ATG

CAT

CAT

SCO

t11,

SCO

t46,

SC

Ot5

0, S

COt5

2,

SCO

t53

XNR_

1595

, XN

R_15

97,

XNR_

2506

, XN

R_28

18,

XNR_

3696

, XN

R_44

68

SVEN

_t12

, SVE

N_t

48,

SVEN

_t54

, SVE

N_

t57

SLI_

1001

1, S

LI_1

0047

, SL

I_10

054,

SL

I_10

056,

SL

I_10

057

SSFG

_RS1

1885

, SS

FG_R

S143

15,

SSFG

_RS1

1875

, SS

FG_R

S183

40,

SSFG

_RS2

3665

SCN

_RS0

7950

, SC

N_R

S151

80,

SCN

_RS1

7555

, SC

N_R

S197

65,

SCN

_RS1

9775

Tyr (

Y)

TAT

ATA

ATA

, GTA

––

––

––

TAC

GTA

GTA

SCO

t48

XNR_

3691

SVEN

_t52

SLI_

1005

2SS

FG_R

S143

40SC

N_R

S175

30

His

(H)

CAT

ATG

ATG

, GTG

––

––

––

CA

CG

TGG

TGSC

Ot1

9XN

R_41

79SV

EN_t

20SL

I_10

019

SSFG

_RS0

0915

SCN

_RS0

9385

Gln

(Q)

CA

ATT

GTT

G–

XNR_

2059

SVEN

_t27

–SS

FG_R

S204

90SC

N_R

S107

85,

CA

GCT

GTT

G, C

TGSC

Ot5

6, S

COt5

9XN

R_12

60, X

NR_

1263

SVEN

_t59

, SVE

N_t

62SL

I_10

059,

SLI

_100

62SS

FG_R

S103

60,

SSFG

_RS1

0345

SCN

_RS2

1830

, SC

N_R

S218

45

Asn

(N)

AAT

ATT

ATT,

GTT

––

––

––

AA

CG

TTG

TTSC

Ot1

2, S

COt1

3XN

R_44

69, X

NR_

4470

SVEN

_t13

, SVE

N_t

14SL

I_10

012,

SLI

_100

13SS

FG_R

S236

55,

SSFG

_RS2

3660

SCN

_RS0

7955

, SC

N_R

S079

60

Page 5 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Tabl

e 1

cont

inue

d

AA

Codo

nA

ntic

odon

Codo

n-

antic

odon

re

cogn

ition

tRN

A g

enes

*

A3(

2)63

**J1

074

65AT

CC 1

0712

65TK

2461

ATCC

146

7268

ATCC

270

6466

12

34

56

78

910

Lys

(К)

AA

ATT

TTT

TSC

Ot4

5XN

R_28

25SV

EN_t

47SL

I_10

046

–SC

N_R

S151

30

AA

GCT

TTT

T, C

TTSC

Ot2

0, S

COt2

1,

SCO

t22

XNR_

1720

, XN

R_17

39,

XNR_

1743

SVEN

_t21

, SVE

N_t

22,

SVEN

_t23

SLI_

1002

0, S

LI_1

0021

, SL

I_10

023

SSFG

_RS2

1980

, SS

FG_R

S218

90,

SSFG

_RS2

1910

SCN

_RS0

9450

, SC

N_R

S095

10,

SCN

_RS0

9530

Asp

(D)

GAT

ATC

ATC

, GTC

––

––

––

GA

CG

TCG

TCSC

Ot4

1, S

COt4

3XN

R_28

32, X

NR_

2835

SVEN

_t43

, SVE

N_t

45SL

I_10

042,

SLI

_100

44SS

FG_R

S182

90,

SSFG

_RS1

8275

SCN

_RS1

5055

, SC

N_R

S150

90

Glu

(E)

GA

ATT

CTT

CSC

Ot4

4XN

R_28

31SV

EN_t

46SL

I_10

045

SSFG

_RS1

8295

SCN

_RS1

5095

GA

GCT

CTT

C, C

TCSC

Ot5

7, S

COt5

8,

SCO

t60

XNR_

1259

, XN

R_12

61,

XNR_

1262

SVEN

_t60

, SVE

N_t

61,

SVEN

_t63

SLI_

1006

0, S

LI_1

0061

, SL

I_10

063

SSFG

_RS1

0340

, SS

FG_R

S103

55,

SSFG

_RS1

0350

SCN

_RS2

1835

, SC

N_R

S218

40,

SCN

_RS2

1850

Cys

(C)

TGT

ACA

AC

A, G

CA

––

––

–

TGC

GCA

GC

Acy

sTXN

R_18

33, X

NR_

5306

SVEN

_t25

, SVE

N_t

5SL

I_10

006

SSFG

_RS2

8225

, SS

FG_R

S214

15SC

N_R

S037

20,

SCN

_RS0

9880

Trp

(W)

TGG

CCA

CCA

SCO

t51

XNR_

3704

SVEN

_t55

SLI_

1005

5SS

FG_R

S142

65SC

N_R

S175

90

Leu

(L)

TTA

TAA

TAA

bldA

XNR_

1995

SVEN

_t26

SLI_

1002

5SS

FG_R

S206

85SC

N_R

S106

10

TTG

CAA

TAA

, CA

ASC

Ot0

9XN

R_48

69SV

EN_t

10SL

I_10

009

SSFG

_RS2

5860

SCN

_RS0

5770

CTT

AAG

AA

G, G

AG

––

––

––

CTC

GAG

AA

G, G

AG

SCO

t08,

SCO

t62

XNR_

0380

, XN

R_51

50SV

EN_t

1, S

VEN

_t66

, SV

EN_t

9SL

I_10

008,

SLI

_100

66SS

FG_R

S055

45,

SSFG

_RS2

7585

SCN

_RS0

4345

, SC

N_R

S249

80

CTA

TAG

AA

G, T

AG

SCO

t23

XNR_

1776

SVEN

_t24

SLI_

1002

2SS

FG_R

S215

85SC

N_R

S096

75

CTG

CAG

TAG

, CA

GSC

Ot3

1XN

R_30

43SV

EN_t

33SL

I_10

032

SSFG

_RS1

7260

SCN

_RS1

4270

Ile (I

)AT

TAA

TA

AT, G

AT–

––

––

–

ATC

GAT

AAT

, GAT

SCO

t33

XNR_

3013

SVEN

_t35

SLI_

1003

4SS

FG_R

S173

70SC

N_R

S140

95

ATA

TAT

AAT

, GAT

, TAT

––

––

––

Page 6 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Tabl

e 1

cont

inue

d

AA

Codo

nA

ntic

odon

Codo

n-

antic

odon

re

cogn

ition

tRN

A g

enes

*

A3(

2)63

**J1

074

65AT

CC 1

0712

65TK

2461

ATCC

146

7268

ATCC

270

6466

12

34

56

78

910

Arg

(R)

CGT

ACG

ACG

, GCG

SCO

t36

XNR_

2898

SVEN

_t38

SLI_

1003

7SS

FG_R

S179

15SC

N_R

S135

75

CGC

GCG

ACG

, GCG

––

––

––

CGA

TCG

ACG

, TCG

––

––

––

CGG

CCG

TCG

, CCG

SCO

t55

XNR_

1483

SVEN

_t58

SLI_

1005

8SS

FG_R

S112

40SC

N_R

S206

05

AG

ATC

TTC

TSC

Ot1

8XN

R_42

58SV

EN_t

19SL

I_10

018

SSFG

_RS2

2445

SCN

_RS0

8865

AG

GCC

TTC

T, C

CT

SCO

t27

XNR_

2176

SVEN

_t29

SLI_

1002

7SS

FG_R

S200

70SC

N_R

S170

25

Gly

(G)

GG

TAC

CA

CC, G

CC–

––

––

–

GG

CG

CCA

CC, G

CCSC

Ot3

9, S

COt4

0,

glyU

aXN

R_28

48, X

NR_

2853

, XN

R_53

07SV

EN_t

4, S

VEN

_t41

, SV

EN_t

42SL

I_10

005,

SLI

_100

40,

SLI_

1004

1SS

FG_R

S181

90,

SSFG

_RS1

8165

, SS

FG_R

S282

30

SCN

_RS0

3715

, SC

N_R

S149

45,

SCN

_RS1

4970

GG

ATC

CA

CC, T

CCSC

Ot1

7XN

R_43

23SV

EN_t

18SL

I_10

017

SSFG

_RS0

2000

, SS

FG_R

S181

95,

SSFG

_RS3

4410

, SS

FG_R

S229

10

SCN

_RS0

8560

, SC

N_R

S149

75

GG

GCC

CTC

C, C

CCSC

Ot3

0XN

R_31

32SV

EN_t

32SL

I_10

031

SSFG

_RS0

6925

, SSF

G_

RS16

450

SCN

_RS1

2860

* St

rain

s ar

e ab

brev

iate

d as

follo

ws:

A3(

2), S

. coe

licol

or A

3(2)

; J10

74, S

. alb

us J1

074;

ATC

C 10

712,

S. v

enez

uela

e AT

CC10

712;

ATC

C146

72, S

. gha

naen

sis A

TCC1

4672

; ATC

C 27

064,

S. c

lavu

liger

us A

TCC2

7064

** To

tal n

umbe

r of t

RNA

gen

es in

gen

ome

Page 7 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

codon AUA. It is not possible for tRNAk2CAU

Ile to recognize codon UUA because of mismatch in second codon position. Hence, there are no tRNA genes in Streptomyces genomes that would allow UUA codon reading (via correct or wobble interactions) in the absence of cognate tRNA

UAALeu. We therefore looked into possibility of UUA mis-

reading. According to Lim and Curran (2001), three anticodons could misread UUA: UAC, GAA, CAA. Of these, first two would lead to aminoacid misincorporation (Val and Phe, respectively).

To gain initial insight into relative mistranslation rate associated with bldA, we applied a computational model of translation accuracy (Shah and Gilchrist 2010) that deduces ratio of abundances of cognate to near-cognate tRNAs (differ from cognate one by one mismatch; see Table 2). The rationale is that error rate would depend not only on the abundance of cognate tRNA, but also on the abundance of all near-cognates, that

Table 2 Gene copy number (GCN) for tRNA genes in six Streptomyces genomes

S. coelicolor S. albus S. venezuelae S. lividans S. ghanaensis S. clavuligerus

tF tN tF tN tF tN tF tN tF tN tF tN

D2

F 1 11 1 12 1 13 1 9 1 12 1 13

Y 1 8 1 9 1 9 1 8 1 9 1 9

H 1 12 1 12 1 13 1 12 1 13 1 12

Q 2 10 3 14 3 14 2 10 3 16 2 15

N 2 12 2 12 2 12 2 12 2 12 2 12

K 4 17 4 19 4 17 4 17 4 18 4 18

D 2 16 2 16 2 17 2 14 2 16 2 14

E 4 15 4 16 4 16 4 15 4 16 4 17

C 1 8 2 8 2 8 1 8 2 8 1 8

D4

A 4 29 4 28 5 28 4 27 5 32 4 30

V 6 28 6 29 6 29 4 28 6 33 4 29

T 4 27 4 27 4 26 4 27 4 27 4 26

P 5 19 4 20 4 22 5 19 5 20 5 20

G 5 22 5 23 5 24 5 20 5 24 5 24

D6

S 4 27 4 28 4 28 4 27 4 28 4 27

L 6 25 6 26 7 24 6 23 7 26 6 26

R 4 19 4 20 4 18 4 19 4 22 4 20

D3

I 1 16 1 17 1 16 1 14 1 15 1 17

D1

M 5 10 6 10 4 10 5 10 4 10 5 10

W 1 6 1 7 1 7 1 6 1 8 1 7

Leu

TAA 1 3 1 3 1 3 1 3 1 3 1 3

CAA 1 10 1 11 1 9 1 10 1 10 1 10

GAG 2 9 2 8 3 8 2 7 3 9 2 9

TAG 1 2 1 3 1 3 1 2 1 3 1 3

CAG 1 6 1 6 1 6 1 6 1 6 1 6

Page 8 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

compete with the former for codon recognition. There was statistically significant posi-tive correlation between the abundance of all leucyl tRNAs and their near-cognates for six Streptomyces species (Fig. 1), suggesting that error rates should not differ for different Leu codon-cognate tRNA pairs (if so, abundances of cognates and near-cognates would be uncorrelated). Similar correlation pattern was observed for most Streptomyces tRNAs (Additional file 1: Fig. S1, S2). We further calculated elongation and error rates for all six leucine codons and revealed that UUA had, in fact, the lowest missense error rate (Table 3). Our findings contrast general belief that low-abundant tRNAs are associated with higher mistranslation rates. Yet, they extend the nuanced view of codon accuracy, based originally on non-actinobacterial, low-GC (less than 70  %) genomes (Shah and Gilchrist 2010), onto GC-rich streptomycetes. Our data also agree with the expectation that proper operation of codon-based genetic switch should be based on accurate trans-lation of UUA.

Rather narrow options for UUA mistranslation, revealed by our analysis, did not take into account that decoding properties of tRNAs can be tuned via posttranscriptional modifications. We identified in genomes of two model streptomycetes a large set of genes for such modifications (including k2C; see above), seven of which are involved in maturation of nascent tRNA

UAALeu in various non-actinomycete bacteria (Table 4 and

Additional file 1: Fig. S3). Of particular interest are the genes for modification of anti-codon loop and adjacent bases of tRNA

UAALeu (see Additional file 1). For example, it is

possible that a posttranscriptional modification of nascent bldA transcript important for UUA decoding and/or tRNA

UAALeu maturation is delayed in Streptomyces. It would tem-

porally limit the occurrence of translationally-competent tRNAUAA

Leu, thus explaining late expression of TTA-containing genes. If so, then streptomycetes deficient in certain tRNA modification genes would resemble bld mutants. We are currently studying this idea using S. albus and S. ghanaensis as experimental models and invite verification of this conjecture for other strains. As a conclusion, our work shows that there are no theo-retical grounds to consider UUA more error prone than the other leucine codons. We examined, in silico, options for UUA mistranslation and draw the attention of research-ers to poorly understood aspects of function of bldA genetic switch.

Fig. 1 Correlation between a focal leucyl tRNA’s abundance tF and the abundance of its neighbors tN, across six Streptomyces genomes (see Additional file 1 for details). Each point represents a leucine tRNA species. The solid lines represent the regression lines between tF and tN for each genome. The data are dependent and nonrandom (Wilcox test, 0.042), and positively correlated (Spearman coefficient, min. 0.354). The mean of the distribution of correlation coefficient values for leucine codons differ from 0 (see Additional file 1: Fig. S2)

Page 9 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Tabl

e 3

Tran

slat

ion

and 

mis

tran

slat

ion

rate

s fo

r six

leuc

ine

codo

ns in

 Str

epto

myc

es

Codo

nCo

gnat

esPs

eudo

-cog

nate

sN

ear-

cogn

ates

S. co

elic

olor

S. a

lbus

S. g

hana

nens

isS.

cla

vulig

erus

S. v

enez

uela

eS.

livi

dans

RcRn

RcRn

RcRn

RcRn

RcRn

RcRn

CU

AU

AG

AA

GC

AG

, GA

G, U

UG

, UCG

, UG

G, U

AA

, UAU

, UA

C2.

067

0.00

72.

067

0.00

92.

067

0.00

92.

067

0.00

92.

067

0.01

02.

067

0.00

7

CU

CG

AG

AA

GU

AG

, CA

G, G

UG

, GCG

, GG

G, G

AA

, GAU

, GA

C4.

134

0.01

44.

134

0.01

44.

134

0.01

24.

134

0.01

46.

201

0.01

24.

134

0.01

1

CU

GC

AG

UA

CA

AG

, UA

G, G

AG

, CU

G, C

CG, C

GG

, CA

A, C

AU, C

AC

2.06

80.

019

2.06

80.

020

2.06

80.

019

2.06

80.

019

2.06

80.

019

2.06

80.

019

CU

UA

AG

GA

G, U

AG

CA

G, A

UG

, ACG

, AG

G, A

AA

, AAU

, AA

C–

––

––

––

––

––

–

UU

AU

AA

AA

AC

AA

, GA

A, U

UA

, UC

A, U

GA

, UAU

, UA

C, U

AG

2.06

70.

006

2.06

70.

006

2.06

70.

006

2.06

70.

006

2.06

70.

006

2.06

70.

006

UU

GC

AA

UA

AA

AA

, GA

A, C

UA

, CC

A, C

GA

, CAU

, CA

C, C

AG

2.06

80.

014

2.06

80.

015

2.06

80.

014

2.06

80.

014

2.06

80.

013

2.06

80.

014

Page 10 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Authors’ contributionsIR generated tN/tF data and carried out computational analysis of codon accuracy; OK analyzed tRNA and tRNA modifi-cation genes in Streptomyces genomes; VF and BO analyzed data; IR and BO drafted the manuscript; all authors read and approved the final manuscript.

AcknowledgementsResearch was supported by the State Fund for Fundamental Research of Ukraine (Project No. F60/2-2015).

Additional file

Additional file 1. Codon correlations and proposed Leu-tRNA modifications.

Table 4 Genes for  tRNA posttranscriptional modification in  S. coelicolor and  S. albus genomes

Protein S. coelicolor A3(2) homolog

S. albus J1074 homolog

Annotation

IscS SCO5486 XNR_1347 tRNAsulfurtransferase, PLP-dependent

IscU SCO1920 XNR_4942 iron-sulfur cluster assembly enzyme

TruA SCO4731 XNR_3758 tRNApseudouridine(38-40) synthase

TruB SCO5709 XNR_1143 tRNApseudouridine synthase B

TruC SCO1625 XNR_4806 tRNApseudouridine synthase C

DusB SCO2497 XNR_4421 tRNA-dihydrouridine synthase B

TrmA SCO5901 XNR_0992 tRNAm5(U34)methyltransferase

TrmB SCO4111 XNR_2813 tRNA (guanine-N(7)-)-methyltransferase

TrmD SCO5594 XNR_1214 tRNA m(1)G37 methyltransferase, SAM-dependent

TrmH SCO4236 XNR_2558 tRNA mG18-2’-O-methyltransferase, SAM-dependent

RluA SCO2073 XNR_4806 23S rRNApseudouridine(746), tRNApseudouridine(32) synthase, SAM-dependent

TadA SCO4038 XNR_2881 tRNA-specific adenosine deaminase

FolE SCO3403 XNR_3431 GTP cyclohydrolase I

QueA SCO1804 XNR_5018 S-adenosylmethionine:tRNAribosyltransferase-isomerase

MnmA SCO5488 XNR_1345 tRNA(Gln,Lys,Glu) U34 2-thiouridylase

MiaA SCO5791 XNR_1074 delta(2)-isopentenylpyrophosphatetRNA-adenosine transferase

MiaB SCO5787 XNR_1078 tRNA-i(6)A37 methylthiotransferase

AroA SCO6819 XNR_1588 5-Enolpyruvylshikimate-3-phosphate synthetase

AroB SCO1494 XNR_5357 3-Dehydroquinate synthase

AroC SCO1496 XNR_5355 Chorismate synthase

AroE SCO1498 XNR_5354 Dehydroshikimatereductase, NAD(P)-binding

AroD SCO1961 XNR_4909 3-Dehydroquinate dehydratase

AroK SCO1495 XNR_5356 Shikimate kinase I

TsaA SCO5032 XNR_4120 tRNA-Thr(GGU) m(6)t(6)A37 methyltransferase, SAM-dependent

TsaB SCO4750 XNR_3789 tRNA(ANN) t(6)A37 threonylcarbamoyladenosine modifi-cation protein; binding partner and protease for TsaD

TsaC SCO5362 XNR_1471 t(6)A37 threonylcarbamoyladenosine biosynthesis protein

TsaD SCO4752 XNR_3791 tRNA(ANN) t(6)A37 threonylcarbamoyladenosine modifi-cation protein; glycation binding protein

TsaE SCO4747 XNR_3786 tRNA(ANN) t(6)A37 threonylcarbamoyladenosine modifi-cation protein; ADP binding protein

TilS SCO3406 XNR_3428 tRNA(Ile)-lysidinesynthetase

Page 11 of 11Rokytskyy et al. SpringerPlus (2016) 5:982

Competing interestsThe authors declare that they have no competing interests.

Received: 31 December 2015 Accepted: 28 June 2016

ReferencesChater KF (2006) Streptomyces inside-out: a new perspective on the bacteria that provide us with antibiotics. Philos Trans

R Soc Lond B Biol Sci 361:761–768. doi:10.1098/rstb.2005.1758Gramajo H, Takano E, Bibb M (1993) Stationary-phase production of the antibiotic actinorhodin in Streptomyces coelicolor

A3(2) is transcriptionally regulated. Mol Microbiol 7(6):837–845. doi:10.1111/j.1365-2958.1993.tb01174.xHackl S, Bechthold A (2015) The gene bldA, a regulator of morphological differentiation and antibiotic production in

Streptomyces. Arch Pharm 348:455–462. doi:10.1002/ardp.201500073Hopwood DA (1987) The Leeuwenhoek lecture. Towards an understanding of gene switching in Streptomyces, the basis

of sporulation and antibiotic production. Proc R Soc Lond B Biol Sci 235:121–138. doi:10.1098/rspb.1988.0067Lawlor E, Baylis H, Chater K (1987) Pleiotropic morphological and antibiotic deficiencies result from mutations in a gene

encoding a tRNA-like product in Streptomyces coelicolor A3(2). Genes Dev 1:1305–1310. doi:10.1101/gad.1.10.1305Lim VI, Curran JF (2001) Analysis of codon: anticodon interactions within the ribosome provides new insights into codon

reading and the genetic code structure. RNA 7:942–957Makitrynskyy R, Ostash B, Tsypik O, Rebets Y, Doud E, Meredith T, Luzhetskyy A, Bechthold A, Walker S, Fedorenko V (2013)

Pleiotropic regulatory genes bldA, adpA and absB are implicated in production of phosphoglycolipid antibiotic moenomycin. Open Biol 3:130121. doi:10.1098/rsob.130121

Marck C, Grosjean H (2002) tRNomics: analysis of tRNA genes from 50 genomes of Eukarya, Archaea, and Bacteria reveals anticodon-sparing strategies and domain-specific features. RNA 8:1189–1232. doi:10.1017/S1355838202022021

Merrick M (1976) A morphological and genetic mapping study of bald colony mutants of Streptomyces coelicolor. J Gen Microbiol 96:299–315. doi:10.1099/00221287-96-2-299

Pettersson BM, Kirsebom LA (2011) tRNA accumulation and suppression of the bldA phenotype during development in Streptomyces coelicolor. Mol Microbiol 79:1602–1614. doi:10.1111/j.1365-2958.2011.07543.x

Shah P, Gilchrist MA (2010) Effect of correlated tRNA abundances on translation errors and evolution of codon usage bias. PLoS Genet 6:e1001128. doi:10.1371/journal.pgen.1001128

Trepanier N, Jensen S, Alexander D, Leskiw B (2002) The positive activator of cephamycin C and clavulanic acid production in Streptomyces clavuligerus is mistranslated in a bldA mutant. Microbiology 148:643–656. doi:10.1099/00221287-148-3-643