RES E AR C H A R T I C L E Open Access

Genome-wide identification and expression

patterns analysis of the RPD3/HDA1 gene

family in cotton

Jingjing Zhang, Aimin Wu, Hengling Wei, Pengbo Hao, Qi Zhang, Miaomiao Tian, Xu Yang, Shuaishuai Cheng,

Xiaokang Fu, Liang Ma, Hantao Wang

and Shuxun Yu

Abstract

Background: Histone deacetylases (HDACs) catalyze histone deacetylation and suppress gene transcription during

various cellular processes. Within the superfamily of HDACs, RPD3/HDA1-type HDACs are the most studied, and it is

reported that RPD3 genes play crucial roles in plant growth and physiological processes. However, there is a lack of

systematic research on the RPD3/HDA1 gene family in cotton.

Results: In this study, genome-wide analysis identified 9, 9, 18, and 18 RPD3 genes in Gossypium raimondii, G.

arboreum, G. hirsutum, and G. barbadense, respectively. This gene family was divided into 4 subfamilies through

phylogenetic analysis. The exon-intron structure and conserved motif analysis revealed high conservation in each

branch of the cotton RPD3 genes. Collinearity analysis indicated that segmental duplication was the primary driving

force during the expansion of the RPD3 gene family in cotton. There was at least one presumed cis-element related to

plant hormones in the promoter regions of all GhRPD3 genes, especially MeJA- and ABA-responsive elements, which

have more members than other hormone-relevant elements. The expression patterns showed that most GhRPD3

genes had relatively high expression levels in floral organs and performed higher expression in early-maturity cotton

compared with late-maturity cotton during flower bud differentiation. In addition, the expression of GhRPD3 genes

could be significantly induced by one or more abiotic stresses as well as exogenous application of MeJA or ABA.

Conclusions: Our findings reveal that GhRPD3 genes may be involved in flower bud differentiation and resistance to

abiotic stresses, which provides a basis for further functional verification of GhRPD3 genes in cotton development and a

foundation for breeding better early-maturity cotton cultivars in the future.

Keywords: Gossypium, Histone deacetylases, Expression patterns, Abiotic stress, Early maturity

Background

DNA combines with nuclear proteins to constitute the

chromatin, which is responsible for storing genetic and

directive information in eukaryotic cells. Chromatin is

highly arranged and mainly composed of nucleosomes,

which are formed by approximately 147 bp of DNA and

an octamer organized by the four core histone proteins_

H3, H4, H2A, and H2B [1]. Gene expression in eukary-

otes involves a complicated interaction, which is con-

trolled not only by the DNA sequence but also by

epigenetic events. Epigenetic mechanisms mainly consist

of histone modification and DNA methylation, and play

an important role in the regulation of gene expression.

In general, histone posttranslational modifications, in-

cluding methylation, acetylation, phosphorylation, ADP-

ribosylation an d ubiquitination, occur at the N-terminal

of histones [2], and these changes facilitate the binding

which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give

appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if

changes were made. The images or other third party material in this article are included in the article's Creative Commons

licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons

licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain

permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the

data made available in this article, unless otherwise stated in a credit line to the data.

* Correspondence: [email protected]; [email protected]

State Key Laboratory of Cotton Biology, Institute of Cotton Research of

Chinese Academy of Agricultural Sciences, Anyang 455000, Henan, China

Zhang et al. BMC Genomics (2020) 21:643

https://doi.org/10.1186/s12864-020-07069-w

of other proteins to DNA, resulting in synergistic or an-

tagonistic regulation of gene transcription [3, 4] . Among

the several histone modifications, histone acetylation is a

reversible process that plays essential roles in epigenetic

regulation. The acetyla tion of core histones is catalyzed

by histone acetyltransferases (HATs) to promote tran-

scriptional activation, whereas deacetylation is regulated

by histone deacetylases (HDACs) that drive the tran-

scriptional suppression [5]. HDACs deacetylate the ly-

sine residues of N-terminal histone tails, resulting in the

repression of gene expression [6].

HDACs are involved in a large amount of biological

processes associated with plant growth and development

[7–9]. Based on sequence homology to yeas t HDACs,

HDACs in plants are divided into three main categories:

reduced potassium dependency 3 / histone deacetylase 1

(RPD3/HDA1), histone deacetylase 2 (HD2), and silent

information regulator 2 (SIR2)[7, 10, 11]. RPD3/HDA1-

type histone deacetylases, which are homologous to

yeast RPD3 and HDA-1, belong to a large family, and

they require zinc ions to catalyze activity; the HDAC in-

hibitor trichostatin A (TSA) or sodium butyrate can in-

hibit their enzymatic activities [7]. The Arabidopsis

RPD3/HDA1 gene family is further classified into three

groups. Class I includes HDA6, HDA7, HDA9, and

HDA19; class II includes HDA5, HDA15, and HDA18;

and HDA2 is the only member of class III [7 , 8]. The

other genes of PRD3/HDA1 family are unclassified in

Arabidopsis.

Over the past 20 years, RPD3/HDA1-type HDACs (call

RPD3 for short below) have been studied extensively as

global regulatory factors playing essential roles in a

series of plant growth and development processes and

the response to various environmental stresses [8, 12–

14]. In Arabidopsis, it has been reported that AtHDA19

was involved in various developmental processes, includ-

ing flowering time, circadian clocks functions, and seed

development [15,

16]. Additionally, AtHDA19 might

regulate gene expression related to jasmonic acid and

ethylene signaling pathways in response to wounding

and path ogen infection [17]. In maize, the expression

patterns of the three ZmPRD3 genes ZmRpd3/101,

ZmRpd3/102, and ZmRpd3/108 showed widespread ex-

pression in all investigated corn organs. Furthermore,

the gene products could be detected in all cellular parts

at specific stages such as kernel, shoot, and anther devel-

opmental periods [18]. In rice, HDA705 responded to

ABA and abiotic stresses, and its expression was induced

by JA. In addition, the expression of HDA702 and

HDA704 was significantly induced by SA, JA, or ABA

[19, 20]. These findings indicate that the RPD3 members

play an important regulatory role in plant development

and in the response to various stresses and plant

hormones.

Cotton is one of the most important economic crops

in China with an essential role in the national economy.

Early maturity and stress resistance are vital target traits

of cotton breeding. Over the past two decades, the RPD3

genes have been intensively studied, and some progress

has been made in Arabidopsis and some other crops.

However, there is a lack of systematic research on the

RPD3 gene family in cotton. Thus, it is necessary to ex-

plore the potential functions of RPD3 genes in cotton. In

our study, the protein seq uences of cotton RPD3-type

HDACs were predicted by genome-wide identification

and the phylogenetic tree, gene structure, conserved

motif, protein do main, expression profiles, and prelimin -

ary functions were comprehensively analyzed. The infor-

mation gained for GhRPD3 provides a reference for

further exploration of the possible functions of RPD3

genes in cotton growth and development.

Results

Identification of RPD3 genes in nine species

In this study, a total of 108 RPD3 protein sequences

from nine species were identified after eliminating re-

dundant sequences, and they are named by the pos-

ition on the chromosome. The corresponding

relationship be tween gene ID n umber and gene name

is shown in Additional file 1:TableS1.Atotalof18

genes (GhHDA1-GhHDA18) containing Hist_deacetyl

(PF00850) domains w ere identified from G. hirsutum;

9 genes were located on the A t genome, and 9 genes

were mapped on the Dt genome. Furthermore, 18

genes (GbHDA1-GbHDA18)fromG. barbadense,9

genes (GaHDA1-GaHDA9)fromG. aboreum,and9

genes ( GrHDA1-GrHDA 9)fromG. raimondii were

detected. Tetraploid c otton possessed twice as many

RPD3 genes as diploid cotton, indicating that no

RPD3 cotton gene was lost in t he process of poly-

ploidy. The numbers of RPD3 genes in the other five

species were 10 (Arabidopsi s), 14 (Oryza sativa L.),

11 (Populus trichocarpa), 8 (Theobroma cacao), and

11 (Zea mays L.). The GhRPD3 protein length ranged

from 232 to 635 aa with an a verage o f 459 aa. The

physicochemical parameters showed that the isoelec-

tricpoint(pl)ofGhRPD3 proteins varied from 4.47

to 8.65 with an average value of 5.68, and the mo-

lecular w eight of GhRPD3 proteins varied f rom 25.79

to 73.01 kDa with an avera ge value of 51.21 k Da. The

subcellular localization results indicated that most of

the GhRPD3 genes were located in cytoplasmic (10)

or nuclear (8), suggesting that GhRPD3 genes might

possess multiple regulatory functions (Table 1). The

predicted length, pI, MW and subcellular localization

of the RPD3 p roteins in other eight species are shown

in Additional file 1: Table S1.

Zhang et al. BMC Genomics (2020) 21:643 Page 2 of 16

Phylogenetic analysis of the RPD3 gene family

A total of 108 identified RPD3 protein sequences from

G. raimondii (9), G. arboreum (9), G. hirsutum (18), G.

barbadense (18), A. thaliana (10), T. cacao (8), Oryza

sativa (14), Zea mays (11) and P. trichocarpa (11) were

employed to construct an unrooted phylogenetic tree

using the neighbor-joining method for investigating the

evolutionary relationships of RPD3 proteins. The RPD3

proteins were phylogenetically classified into 4 subfam-

ilies (Class I, Class II, Class III, and unclassified) accord-

ing to the formulated subfamilies in Arabidopsis [7]. The

Class I subgroup was the largest subfamily with 49

RPD3 genes, whereas the Class III subgroup has the few-

est members, only containing one gene in seven diploid

species genomes and two genes in two tetraploid cotton

genomes (Fig. 1). Among these four classes, each sub-

family contained RPD3 genes from all nine species, indi-

cating this gene family was relatively conserved in

different species during evolution.

Exon-intron structure and conserved motif analysis

The domains of the RPD3 sequences in cotton were in-

vestigated and exhibited according to the results of the

SMART database using TBtools, revealing that all cotton

RPD3 genes contained a Hist_deacetyl domain (Add-

itional file 2: Table S2 and Additional file 3: Figure S1).

An unrooted phylogenetic tree with the predicted cotton

RPD3 genes was constructed (Fig. 2a), and then exon-

intron structure (Fig. 2b) and con served motifs (Fig. 2c)

were analyzed to better understand the similarity and

differences of cotton RPD3 members. The results

showed that the len gth of RPD3 cotton genes was rela-

tively conserved in Class I and Class III, but there were

twelve longer sequences in Class II and the unclassified

group. The RPD3 cotton genes included from 3 to 17

exons and most RPD3 genes (48/54) contained more

than five exons (Additional file 4: Table S3), which

might be associated with the diversification of their

functions. In terms of the distribution of motifs, most

RPD3 cotton genes belonging to the same subfamily

showed a similar motif composition, except in the un-

classified group (Fig. 2c). Most Class I subfamily mem-

bers contained 9 motifs, whereas GrHDA5 and GhHDA4

contained 4 and 6 motifs, respectively. Class III subfam-

ily genes had three or four motifs, and most Class II sub-

family members possessed 7 motifs, except for

GhHDA12 with 6 motifs. There were differences in the

exon-intron structure and motif arrangement among the

four categories, but they were highly conserved on the

same branches, indicati ng that the RPD3 members clas-

sified into the same branch might perform a relatively

conserved function in cotton growth and development.

Chromosomal distribution, gene duplication and selection

pressure

The chromosomal distributions of GrRPD3 , GaRPD3,

GbRPD3, and GhRPD3 genes were visualized according

to the genomic position of 54 cotton RPD3 genes (Add-

itional file 5: Table S4 and Fig. 3). In G. hirsutum,18

GhRPD3 genes were unevenly mapped on 13

Table 1 Physicochemical parameters of 18 RPD3 genes in G. hirsutum

Name Gnen ID Protein Length Protein pI Protein MW (kD) Subcellular localization

GhHDA1 Ghir_A01G001410.1 499 4.9676 56.18 Nuclear

GhHDA2 Ghir_A03G007210.1 471 5.076 53.09 Nuclear

GhHDA3 Ghir_A03G008200.1 655 5.325 73.01 Cytoplasmic

GhHDA4 Ghir_A03G018610.1 351 4.4737 39.55 Cytoplasmic/Nuclear

GhHDA5 Ghir_A05G039610.1 449 6.9085 48.66 Mitochondrial/Chloroplast

GhHDA6 Ghir_A09G010210.1 429 4.8969 49.08 Cytoplasmic/Nuclear

GhHDA7 Ghir_A12G027820.1 574 6.3108 63.26 Cytoplasmic

GhHDA8 Ghir_A13G019980.1 232 6.5919 25.79 Plasma Membrane

GhHDA9 Ghir_A13G023460.1 368 5.3373 40.37 Cytoplasmic/Chloroplast

GhHDA10 Ghir_D01G001410.1 499 4.9676 56.26 Nuclear

GhHDA11 Ghir_D02G019970.1 465 5.1309 52.65 Nuclear

GhHDA12 Ghir_D03G010660.1 635 4.8889 71.02 Cytoplasmic

GhHDA13 Ghir_D03G011510.1 471 5.1489 53.06 Nuclear

GhHDA14 Ghir_D04G003510.1 443 6.9591 47.95 Chloroplast/Mitochondrial

GhHDA15 Ghir_D09G009940.1 429 4.8371 49.11 Cytoplasmic/Nuclear

GhHDA16 Ghir_D12G027930.1 579 6.1788 63.80 Cytoplasmic

GhHDA17 Ghir_D13G020760.1 331 8.6517 37.28 Plasma Membrane

GhHDA18 Ghir_D13G024090.1 380 5.648 41.63 Cytoplasmic

Zhang et al. BMC Genomics (2020) 21:643 Page 3 of 16

chromosomes. A03 contained the most GhRPD3

genes (3), whereas the other 12 chromoso mes only

contained one or two GhRPD3 genes (Fig. 3a). The

chrom osomal distribution of 18 GbRPD3 genes in G.

barbadense was similar to that of GhRPD3 gene s in

G. hirsutum (Fig. 3b). In G. arboreum,9GaRPD3

genes were unevenly located on 6 chromosomes.

Chr01 and Chr13 contained three and two GaRPD3

genes, respectively, and the other 4 chromosomes

contained only one GaRPD3 gene (F ig. 3c). In G. rai-

mondii, the chromosomal distribution of 9 GrRPD3

genes was highly consistent with the corresponding D

subgenome of G. hirsutum (Fig. 3d), sh owing con-

served numbers a nd chromosomal distribution of

RPD3 genes between diploid and tet raploid cotton

species. In addition, the lopsided chromosomal distri-

bution of the cotton RPD3 genes indicated that

genetic variation occurred during evolution. Notably,

most of the RPD3 genes were distributed on the o p-

posite ends of the chromosomes in f our cotton spe-

cies (Fig. 3).

In general, tandem and segmental duplication are

two of the main reasons for gene family generation

during evolution [21].Theanalysisofgeneduplica-

tion indicated that all RPD3 family members were

amplified o nly through segmental duplication (Add-

itional file 6: Table S5), suggesting that segmental du-

plication played a vital role in the evolution of the

RPD3 gen e family. The homologous gene pairs ob -

tained by collinearity analysis among RPD3 genes in

G. arboreum, G. raim ondii,andG. hirsutum were vi-

sualized using circular maps (Fig. 4). The Ka/Ks ratios

of most homologous gene pairs were lower than one,

indicating that purified selection was essential during

Fig. 1 Neighbor-joining phylogenetic tree of RPD3 gene family. The 108 predicted RPD3 proteins from G. hirsutum, G. arboreum, G. barbadense, G.

raimondii, A. thaliana, P. trichocarpa, T. cacao, Oryza sativa, and Zea mays were aligned using ClustalW, and the neighbor-joining (NJ) method was

used to construct this unrooted phylogenetic tree using MEGA 7.0 program with 1000 bootstrap repetitions. Four subfamilies are represented by

the different colored lines

Zhang et al. BMC Genomics (2020) 21:643 Page 4 of 16

the evolution of cotton RPD3 genes, whereas the Ka/Ks

ratios of two gene pairs (GhHDA2/GaHDA3 and

GhHDA6/GaHDA6)weremorethan1,suggestingthat

these two pairs might have experienced positive selection

pressure. The study also predicted the occurrence time of

segmentally duplicated RPD3 gene pairs by the formula

“t = Ks/2r” (r =2.6X10

− 9

)[22]. Except for the GhHDA6/

GaHDA6 gene pair, the other segmental duplication

events of three cotton species might have occurred 0.6 to

144.44 million years ago (MYA) with an average time of

18.39 million years ago (Additional file 6: Table S5).

Analysis of cis-elements in predicted promoter regions of

GhRPD3

To explore the possible regulatory functions of GhRPD3

genes under various envir onmental stresses and hor-

mone regulation pathways, the 2000-bp promoter re-

gions of 18 GhRPD3 genes were submitted to the

PlantCARE database for the identification of putative

stress-associated and plant hormone-related cis-

elements. A total of 9 kinds of elements related to plant

hormones, containing AuxRE-core (auxin), TGA-

element (auxin), P-box (gibberellin), TATC-box

Fig. 2 Phylogenetic relationships, exon-intron structure, and conversed motif analysis of cotton RPD3 genes. a A neighbor-joining phylogenetic

tree of 54 cotton RPD3 genes was generated using the MEGA7.0 program; (b) Exon-intron structure analysis of 54 cotton RPD3 genes. The UTRs,

exons, and introns are represented with yellow boxes, green boxes, and black lines, respectively; (c) The 10 conversed protein motifs of RPD3

genes are indicated by different colored boxes

Zhang et al. BMC Genomics (2020) 21:643 Page 5 of 16

(gibberellin), GARE- motif (gibberellin), CGTAC -motif

(MeJA), TGACG-motif (MeJA ), TCA-element (SA), and

ABRE (ABA), and 4 kinds of elements responding to

stresses, including TC-rich repeats (defense and stress

responsiveness), MBS (drought), WUN-motif (wound)

and LTR (cold stress), were predicted in the promoters

of GhRPD3 genes. As shown in Fig. 5, the promoters of

some GhRPD3 genes contained various hormone-

responsive and stress-responsive components, such as

GhHDA2 (2 MBS, 2 LTR, 2 TC-rich repeats, 1 GARE-

motif, 2 ABRE, 1 TGACG-motif) and GhHDA13 (1

MBS, 1 LTR, 1 TC-rich repeats, 1 AuxRE-core, 2 GARE-

motif, 1 TCA-element, 4 ABRE, 2 TGACG-motif).

Among the 18 GhRPD3 genes, there are large numbers

of light-responsive elements distributed in their pro-

moter regions (Additional file 7: Table S6). In addition,

MeJA-responsive and ABA-responsive elements are

more common than other hormone-related elements

(Additional file 8: Figure S2). These results revealed that

GhRPD3 genes might be involved in MeJA and ABA

hormone signaling pathways as well as response to en-

vironmental stresses.

Expression profiles of GhRPD3 genes in different tissues

and under different abiotic stresses

To understand the potential functions of GhPRD3 genes in

the growth and development of cotton, we studied their

expression in various cotton tissues, including the anther,

pistil, bract, sepal, petal, filament, torus, root, leaf, stem,

ovules, and fibers, using publicly available transcriptomic

data provided by Hu et al. [23]. Transcripts of all the

GhRPD3 genes were detected in at least three tissues with

fragments per kilobase million (FPKM) ≥ 1. Furthermore,

ten genes exhibited high expression levels in all selected tis-

sues (Additional file 9: Table S7). These results indicated

that GhRPD3 genes are widely expressed in both repro-

ductive organs and vegetative organs and thus might have

multiple biological functions. After log2-conversion of

FPKM values, the expression profiles of GhRPD3 genes in

differen t tissues are shown in Fig. 6a. Seven GhRPD3 genes

exhibited relatively high expression levels in at least eight

tissues (log2-transformed FPKM value≥2.6); in particular,

one pair of homologous genes (GhHDA1/GhHDA10)

showed a high expression level in all the tissues with a simi-

lar expression pattern. Nevertheless, three GhRPD3 genes

(GhHDA4, GhHDA14, GhHDA18) were expressed at rela-

tively low levels in at least eight tissues (log2-transformed

FPKM value< 1), of which GhHDA14 showed the lower

expression in all tissues except for the pistil. These

homologous gene pairs (GhHDA1/Gh HDA 10, GhHDA4/

GhHDA11, GhHDA2/GhHDA13, GhHDA6/GhHDA15,

GhHDA7/GhHDA16,andGhHDA9/GhHDA18)werelo-

cated on At and Dt subgenomes and exhibited similar ex-

pression patterns. For example, homologous gene pairs

Fig. 3 Chromosomal distribution of cotton RPD3 genes. a, b, c and d represent the chromosomal location of RPD3 genes from G. hirsutum (a), G.

barbadense (b), G. arboreum (c), and G. raimondii (d), respectively. The chromosome number is shown on the top of each chromosome. The scale

bars represent the length in mega bases (Mb)

Zhang et al. BMC Genomics (2020) 21:643 Page 6 of 16

(GhHDA4/GhHDA11 and GhHDA9/GhHDA18)showed

relative ly low expression in all twelve tissues. The

gene pair GhHDA2/GhHDA13 exhibited relatively

high expres sion in torus and ovule but relatively low

expression in petals (Fig. 6a).

Based on the analysis of cis-elements in promoter re-

gions and previous reports on RPD3 genes in other

plants, GhRPD3 gens might respond to abiotic stresses.

To test this hypothesis, we investigated the expression

characteristics of 18 GhRPD3 genes under cold, heat,

PEG, and salt treatments using available transcriptomic

data [23] (Fig. 6b). The expression of most GhRPD3

genes were induced by the four stresses to varying de-

grees. GhHDA1, GhHDA2, GhHDA6, GhHDA10,

GhHDA12, and GhHDA18 showed upregulated expres-

sion under four stress treatments. However, one gene

(GhHDA4) exhibited marked downregulation in the

presence of the four abiotic stresses. Some genes can re-

spond to one specific abiotic stress. For example, the ex-

pression of GhHDA13 and GhHDA16 was significantly

induced by PEG treatment. Four genes (GhHDA7,

GhHDA11, GhHDA5) showed upregulated expression

Fig. 4 RPD3 homologous gene pairs among G. arboreum, G. raimondii and G. hirsutum. Orange, blue and red represent chromosomes of G.

arboreum, G. raimondii and G. hirsutum, respectively

Zhang et al. BMC Genomics (2020) 21:643 Page 7 of 16

under heat treatment. The expression of GhHDA9 was

significantly upregulated under cold and salt treatments.

According to the results, we can conclude that GhRPD3

genes play an essential role in response to abiotic

stresses.

Characterization of GhRPD3 genes expression during

flower bud differentiation

To explore expression differences of GhRPD3 genes be-

tween early-maturity and late-maturity cottons during

flower bud differentiation, we selected nine genes show -

ing relatively high expression in floral organ tissues

(anther, pistil, bract, sepal, petal, filament and torus) for

qRT-PCR. The buds of an early-maturity variety

(CCRI50) and a late-maturity variety (GX11) from the

one-leaf to five-leaf stage were used for qRT-PCR (Fig. 7).

The results revealed that more than half of these genes

(5/9) possessed relatively higher expression in early-

maturity cotton compared with late-maturity cotton

during flower bud differentiation. GhHDA5 showed

marked differences at the two-leaf and three-leaf stages,

and these two stages were regarded as the important

period of flower bud differentiation. A homologous gene

pair (GhHDA6/GhHDA15) located on At and Dt

Fig. 5 Cis-elements of GhRPD3 genes in promoter regions. The numbers of different cis-elements are presented in the form of bar graphs, and

similar cis-elements are exhibited with the same colors

Fig. 6 Expression patterns of RPD3 genes in G. hirsutum. a and b represent the expression patterns of GhRPD3 genes in different tissues ( a) and

under four different abiotic stresses (b), respectively. Gene names are shown on the right. Scale bars on the right represent the log2-transformed

FPKM values of each gene

Zhang et al. BMC Genomics (2020) 21:643 Page 8 of 16

respectively, showed the same expression trend. Both of

them presented the highest expression at three-leaf stage

and then exhibited downregulated expression in next

two stages in CCRI50. In addition, all nine genes showed

relatively higher expression at the two-leaf or three-leaf

stage in CCRI50 compared with GX11. The results

showed that GhRPD3 genes are associated with the early

maturity of cotton.

Responses of GhRPD3 genes to MeJA and ABA treatment

MeJA and ABA play imp ortant roles in plant stress re-

sistance. To further explore the possible functions of

GhRPD3 genes, we selected the GhRPD3 genes

containing MeJA- and ABA-responsive elements in the

predicted promotors to analyze their expression charac-

teristics under MeJA and ABA treatment by qRT-PCR

(Figs. 8 and 9). Most GhRPD3 genes (8/13) were mark-

edly upregulated at 9 h after MeJA treatment. Three

genes (GhHDA7, GhHDA13,andGhHDA18)exhibited

significantly upregulated expression at three or more time

points, whereas four genes (GhHDA2, GhHDA8,

GhHDA9,andGhHDA11)showedmarkedtranscriptional

downregulation at least three time points after MeJA

treatment (Fig. 8). More than half of the GhRPD3 genes

(6/11) were significantly upregulated at 9 h after ABA

treatment. Three GhRPD3 genes (GhHDA14, GhHDA15,

and GhHDA18) showed relatively high expression at three

or more time points, whereas three GhRPD3 genes

(GhHDA10, GhHDA11,andGhHDA17)showedearly

downregulated and then upregulated expression patterns

under ABA treatment (Fig. 9). The results showed that the

exogenous application of MeJA and ABA significantly in-

duced the transcription of most GhRPD3 genes containing

MeJA-responsive and ABRE elements in their promoter

regions.

Discussion

Among the several histone modifications, histone acetyl-

ation plays an essential role in plant growth and devel-

opment [24]. Histone acetylation and deacetylation are

catalyzed by histone acetyltransferases (HATs) and his-

tone deacetylases (HDACs), respectively [20]. In plants,

HDACs are involved in a variety of biological processes

associated with plant growth and development [25].

Fig. 7 Expression levels of 9 GhRPD3 genes between CCRI50 and GX11. Blue and orange bar graphs indicate the expression of early-maturity

cotton (CCRI 50) and late-maturity cotton (GX11), respectively. The error bars show the standard deviation of three biological replicates

Zhang et al. BMC Genomics (2020) 21:643 Page 9 of 16

Fig. 8 Expression profiles of 13 GhRPD3 genes under MeJA treatment. Orange boxes represent the MeJA-responsive elements of 13 GhRPD3

genes in the promoter regions (left). The expression changes of 13 GhRPD3 genes under MeJA treatment are shown using a heatmap (right).

qRT-PCR was carried out with three technical and three biological replicates. Relative expression levels of each gene were calculated after

normalizing the expression level in CK (water) to 1.0

Fig. 9 Expression patterns of 11 GhRPD3 genes under ABA treatment. Green boxes represent the ABRE of 11 GhRPD3 genes in the promoter

regions (left). The expression changes of 11 GhRPD3 genes under ABA treatment are shown using a heatmap (right). qRT-PCR was conducted

with three technical and three biological replicates. Relative expression levels of each gene were calculated after normalizing the expression level

in CK (water) to 1.0

Zhang et al. BMC Genomics (2020) 21:643 Page 10 of 16