Comprehensive Genome-Wide Analysis of Histone Acetylation Genes in Roses and Expression Analyses in Response to Heat Stress

Roses have high economic values as garden plants and for cut-flower and cosmetics industries. The growth and development of rose plants is affected by exposure to high temperature. Histone acetylation plays an important role in plant development and responses to various stresses. It is a dynamic and reversible process mediated by histone deacetylases (HDAC) and histone acetyltransferases (HAT). However, information on HDAC and HAT genes of roses is scarce. Here, 23 HDAC genes and 10 HAT genes were identified in the Rosa chinensis ‘Old Blush’ genome. Their gene structures, conserved motifs, physicochemical properties, phylogeny, and synteny were assessed. Analyses of the expression of HDAC and HAT genes using available RNAseq data showed that these genes exhibit different expression patterns in different organs of the three analyzed rose cultivars. After heat stress, while the expression of most HDAC genes tend to be down-regulated, that of HAT genes was up-regulated when rose plants were grown at high-temperature conditions. These data suggest that rose likely respond to high-temperature exposure via modification in histone acetylation, and, thus, paves the way to more studies in order to elucidate in roses the molecular mechanisms underlying rose plants development and flowering.


Introduction
Histone acetylation is well known to be involved in different processes of plant growth and development [1][2][3], and in response to environmental changes [4], such as heat stress [5]. Histone acetylation has become an important part of epigenetics and has received considerable attention recently. Histone acetylation/deacetylation is a dynamic and reversible process mediated by HATs and HDACs. HDACs are involved in histones deacetylation, inhibiting transcription of functional genes, and, ultimately, silencing their expression [6,7]. In Arabidopsis thaliana, these HDACs can be divided into three subfamilies: RPD3/HDA1, HD2, and SIR2 [8]. Members of the HD2 subfamily are plant-specific HDACs [9]. HATs can neutralize the positive charge on lysine residues, weaken the binding effect of histone to DNA, induce relaxed chromatin structure, and facilitate the binding of transcription factors or transcriptional regulatory proteins to DNA, thus promoting gene transcription [10,11]. HATs can be divided into four subfamilies, named GNAT, MYST, CBP, and TAF II 250 [8]. Although a number of studies, primarily using the plant model

The Identification of HDACs and HATs
HDAC and HAT gene members were retrieved in the six species (R. chinensis, R. rugosa, R. multiflora, F. vesca, A. thaliana, and Oryza sativa; Table 1) based on previous reports or by BLAST search.
In F. vesca, alternative splicing transcript variants were identified for a number of HDAC and HAT genes, all of which retained the t1 suffix for further analyses. The number of HDAC and HAT genes varied among the six species (Table 1). Blastp and conserved domain search identified 23 HDACs in R. chinensis, 26 HDACs in R. multiflora, 12 HDACs in R. rugosa and 14 HDACs in F. vesca. Concerning the HAT genes, 10 were identified in R. chinensis, 15 in R. multiflora, 11 in R. rugosa, and 13 in F. vesca (Tables S3-S6). They were named according to the naming conventions in Arabidopsis [8].
Compared with Arabidopsis, R. chinensis has a higher number of HDAC genes, and lower number of HAT genes (Table 1). In addition, the number of RPD3/HDA1 subgroup members varied greatly between the rose and F. vesca, with 19 members in R. chinensis and only 10 members in F. vesca. Only 10 RPD3/HDA1 subgroup members were found in R. rugosa. Similarly, while six members of HD2 subfamily were identified in R. multiflora, only two members were identified in R. chinensis and only one member was identified in R. rugosa. The differences in HDAC and HAT gene numbers (Table 1) could be due to the possibility that these rose species may have evolved independently regarding the process of histone acetylation. This is supported for example by the recent report that proposed that R. rugosa diverged from R. chinensis and over 6 million years ago [35].

Gene Structure and Conserved Domains
Structure analyses of coding sequences showed that HDAC genes have different numbers of exons and introns. In R. chinensis, the SIR2 subfamily genes have high number of introns, while the HD2 subfamily genes have fewer introns (Figure 1). The number of introns in genes of R. chinensis also differed from that of A. thaliana and F. vesca (Figures 1 and S1). Genes of the RPD3/HDA1 subfamily had the highest number of introns, reaching with seventeen introns in the case of RcHDA15, although some members have no or only one or two introns. HD2 subfamily genes of A. thaliana and F. vesca have higher number of introns compared to R. chinensis (Figures 1 and S1), while the number of introns in RPD3/HDA1 and SIR2 subfamilies were similar in R. chinensis, in A. thaliana, and in F. vesca, with the highest number of introns reaching 17 in RcHDA15, AtHDA15, and FvHDA15. To some extent, this indicated that the RPD3/HDA1 subfamily and SIR2 subfamily were conserved among the studied species here, while the number of introns in the HD2 subfamily of rose was different from that of Arabidopsis and strawberry. The number of exons and introns also varied between HAT genes of R. chinensis, of A. thaliana and of F. vesca. (Figures S2 and S3). For example, RcHAC4, RcHAF2, FvHAC4, and FvHAF2 had fewer introns than AtHAC4 and AtHAF2. one or two introns. HD2 subfamily genes of A. thaliana and F. vesca have higher number of introns compared to R. chinensis (Figures 1 and S1), while the number of introns in RPD3/HDA1 and SIR2 subfamilies were similar in R. chinensis, in A. thaliana, and in F. vesca, with the highest number of introns reaching 17 in RcHDA15, AtHDA15, and FvHDA15. To some extent, this indicated that the RPD3/HDA1 subfamily and SIR2 subfamily were conserved among the studied species here, while the number of introns in the HD2 subfamily of rose was different from that of Arabidopsis and strawberry. The number of exons and introns also varied between HAT genes of R. chinensis, of A. thaliana and of F. vesca. (Figures S2 and S3). For example, RcHAC4, RcHAF2, FvHAC4, and FvHAF2 had fewer introns than AtHAC4 and AtHAF2.
The conserved domains in the 23 RcHDACs and in the 10 RcHATs were analyzed by NCBI-CDD ( Figure S4). All members of the RPD3/HDA1 subfamily possess the characteristic domains of HDAC family. The two members of the SIR2 subfamily, RcSRT1 and RcSRT2, contain the conserved domains SIRT4 or SIRT7, respectively. HD2 members have a highly conserved NPL domain, whereas the GNAT subfamily proteins RcHAG1, RcHAG2, and RcHAG3 (includes RcHAG3-1 and RcHAG3-2) have 3 different conserved domains, COG5076, Hat1_N and ELP3, respectively. RcHAG4 have the conserved domain of MYST, designated PLN00104. The members of CBP all have a conserved HAT_KAT11 domain, and TAF II 250 members have a highly conserved DUF3591 domain.

Physicochemical Properties of HDACs and HATs
Expasy was used to analyze the physicochemical properties of RcHDACs and RcHATs ( Table 2). The size of R. chinensis HDAC proteins varied greatly, with the largest protein composed of 661 amino acids residues and the smallest composed of 46 amino acids residuces. The theoretical PI of SIR2 subfamily is about 9. The theoretical PI of RPD3/HDA1 and HD2 subfamily members were discrepant, and there were great differences among subfamily members. In terms of protein stability, SIR2 subfamily proteins are predicted to be unstable proteins, and HD2 subfamily proteins are predicted as stable proteins. In total, 11 members of the RPD3/HDA1 subfamily are predicted to be stable proteins (i.e., RcHDA14-2), and 8 are predicted to be unstable proteins (ie. RcHDA2-6). SIR2 subfamily proteins were all hydrophilic, while members of the HD2 and RPD3/HDA1 subfamilies were more hydrophobic. Hydrophilic proteins such as RcHDA6-2 and RcHDA19 were also present in the RPD3/HDA1 subfamily. RcHATs proteins were all hydrophilic. The size of RcHAT proteins varied greatly, with the largest protein containing 1900 amino acid residues and the smallest protein containing 95 amino acid residues. All HAT proteins, except RcHAG3-1 and RcHAG3-2, were rated as likely unstable proteins.

RPD3/HDA1 Subfamily
To know more about the phylogenetic relationships among RPD3/HDA1 genes, a phylogenetic analysis of RPD3/HDA1 proteins from R. chinensis (Table S3), from A. thaliana (Table S1) and from F. vesca (Table S5) was constructed by NJ method. RcRPD3/HDA1 proteins and FvRPD3/HDA1 proteins were divided into 4 clades named Class I to Class IV with reference to the classification of the A. thaliana (Figure 2a). While R. chinensis, F. vesca and A. thaliana have comparable numbers of gene member for Class I and Class II, R. chinensis has more gene members for Class III (8 genes for R. chinensis to one in F. vesca and one in A. thaliana) and Class IV (5 genes for R. chinensis, 2 in F. vesca and 2 in A. thaliana).

HD2 Subfamily
Phylogenetic analysis of R. chinensis HD2 subfamily proteins was constructed with F. vesca and A. thaliana HD2 proteins by NJ method (Figure 2b). RcHDT1 and RcHDT2 are more closely related to FvHDT3-1 and FvHDT3-2, respectively, than to the HD2 proteins of A. thaliana. The number of HD2 subfamily proteins varied among species, with 4 members in A. thaliana, and 2 members in R. chinensis and F. vesca.

HD2 Subfamily
Phylogenetic analysis of R. chinensis HD2 subfamily proteins was constructed with F. vesca and A. thaliana HD2 proteins by NJ method (Figure 2b). RcHDT1 and RcHDT2 are more closely related to FvHDT3-1 and FvHDT3-2, respectively, than to the HD2 proteins of A. thaliana. The number of HD2 subfamily proteins varied among species, with 4 members in A. thaliana, and 2 members in R. chinensis and F. vesca.

SIR2 Subfamily
R. chinensis, F. vesca, and A. thaliana have two members each of SIR2 protein. Phylogenetic analysis shows that the SIR2 subfamily is highly conserved among the 3 species (Figure 2c).

HAT Family
Phylogenetic analysis divided HAT proteins into four subfamilies: MYST, CBP, TAF II 250, and GNAT ( Figure 2d). The GNAT subfamily was previously divided into four classes designated as GCN5, ELP3, HAT1, and HPA2 [8]. In Arabidopsis, the GCN5, ELP3, and HAT1 classes are each composed of a single gene (AtHAG1, AtHAG3, and AtHAG2, respectively), whereas no members of the HPA2 class have been identified in A. thaliana [8].

Chromosomal Location, Homologous Genes, and Synteny
The 23 HDAC and 10 HAT genes are distributed on the 7 chromosomes of the rose (Figure 3a).
Synteny analysis with R. rugosa and F. vesca, two closely related species to R. chinensis, revealed that homologs of 10 HDAC genes were found in F. vesca genome and nine in R. rugosa genome ( Figure 3b; indicated with orange lines). Nine HAT homologous genes were found in F. vesca and in R. rugosa genomes (Figure 3b, blue lines). The remaining members of HDACs, such as RcHDA14-3, RcHDA2-2, and RcHDA2-3 found on chromosomes 1 and 4, could not be found in R. rugosa and F. vesca genomes (Figure 3b). It is possible that these HDACs are unique to Rosa compared to Fragaria. The absence of these HDACs in the R. rugosa genome is likely related to the fact that the available genome sequence of R. rugosa is not be complete to identify these genes. Synteny analysis with R. rugosa and F. vesca, two closely related species to R. chinensis, revealed that homologs of 10 HDAC genes were found in F. vesca genome and nine in R. rugosa genome (Figure 3b; indicated with orange lines). Nine HAT homologous genes were found in F. vesca and in R. rugosa genomes (Figure 3b, blue lines). The remaining members of HDACs, such as RcHDA14-3, RcHDA2-2, and RcHDA2-3 found on chromosomes 1 and 4, could not be found in R. rugosa and F. vesca genomes (Figure 3b). It is

Expression of HDACs and HATs in Different Organs of Three Roses
The expression patterns of HDACs and HATs in various organs of 2 Rosa species (R. chinensis, R. wichurana) and 1 modern rose cultivar (R. hybrida 'Yesterday') were analyzed. Expression analysis using published RNAseq data [30,31] shows that RcHDA6-1, RcHDA2-6, RcHDA2-7, RcHDA2-8, RcHDA14-2, RcHDA14-3, and RcHDA14-4 are not expressed in all tested tissues of the three roses ( Figure 4). HDAC genes have different expression levels depending on the organ studied. For example, RcHDA6-2, RcHDA14-1, and RcSRT2 are highly expressed in roots, leaves, and pink petals of R. chinensis 'OB', respectively (Figure 4a). Whereas high transcript abundance for RcHDA19, RcHDT1, and RcHDT2 were observed in green petals of R. chinensis 'OB'. In 'Rw', RcHDA19, RcHDT1, and RcHDT2 were highly expressed in flower buds, a stage where petals are still green, thus similar to 'OB'. RcHDA6-2 showed the highest transcript abundance in stems of 'Rw' and 'Ry' roses, while RcHDA14-1 was highly expressed in leaves (Figure 4b). These data shows HDAC genes expressed are expressed in different tissues and organs of the rose, and suggest their potential roles in vegetative and reproductive growth.  According the RNAseq data [30,31] HAT genes also show different expression levels in different tissues of the rose. The 2 HAT genes RcHAG3-2 and RcHAC4 are likely not expressed in roots and stems of Rw' and 'Ry' roses ( Figure 5), while RcHAG3-1 is highly expressed in developing petals at onst of color change in 'OB' flower buds (Figure 5a). RcHAG2 showed the highest transcript abundance in flower buds of 'Rw' rose, respectively (Figure 5b, Table S7), and transcript of RcHAC2 highly accumulated in roots of 'Rw' and 'Ry' roses. The fact that HAT genes are expressed in different tissues of the rose suggest their likely roles in histone acetylation during plant organs development.
Genes 2022, 13, x FOR PEER REVIEW 12 of 20 respectively (Figure 5b, Table S7), and transcript of RcHAC2 highly accumulated in roots of 'Rw' and 'Ry' roses. The fact that HAT genes are expressed in different tissues of the rose suggest their likely roles in histone acetylation during plant organs development.
qRT-PCR analyses showed that in 'CL' roses, the class I gene RcHDA6-1 was highly expressed in leaves and pistils of the control plants, and following high-temperature treatments, its expression was significantly decreased in these organs, while its expression was increased in stamens ( Figure 6; Table S8). The expression of RcHDA15 (Class II) and RcHDA2-2(Class III) also decreased in leaves and sepals in response to high-temperature stress. RcHDA2-1(Class III), RcHDA2-5(Class III), RcHDA8(Class IV), and RcHDA14-2 (Class IV) also decreased after high-temperature treatments in leaves, pistils, and sepals. RcSTR2 (SIR2 subfamily) was expressed in the leaves, and its expression level also decreased after high-temperature treatment. HD2 subfamily members, RcHDT1 and RcHDT2, were expressed in almost all tissues analyzed, with the highest expressed level in the leaves or stamen, respectively (Table S8). Their expression also decreased after hightemperature treatment. Genes 2022, 13, x FOR PEER REVIEW 13 of 20  qRT-PCR analyses showed that in 'CL' roses, the class I gene RcHDA6-1 was highly expressed in leaves and pistils of the control plants, and following high-temperature treatments, its expression was significantly decreased in these organs, while its expression was increased in stamens ( Figure 6; Table S8). The expression of RcHDA15 (Class II) and RcHDA2-2(Class III) also decreased in leaves and sepals in response to high-temperature stress. RcHDA2-1(Class III), RcHDA2-5(Class III), RcHDA8(Class IV), and RcHDA14-2 (Class IV) also decreased after high-temperature treatments in leaves, pistils, and sepals. RcSTR2 (SIR2 subfamily) was expressed in the leaves, and its expression level also decreased after high-temperature treatment. HD2 subfamily members, RcHDT1 and RcHDT2, were expressed in almost all tissues analyzed, with the highest expressed level in the leaves or stamen, respectively (Table S8). Their expression also decreased after hightemperature treatment.
In contrast to HDAC genes, the expression of HAT genes RcHAG1, RcHAG2, and RcHAF2 increased after high-temperature treatments in stamens, pistils, and leaves (Figure 6b, Table S8). Similarly, the expression of RcHAC4 was also up-regulated in petals, stamens, and leaves after high-temperature treatments ( Figure 6; Table S8). RcHAG4 was expressed in a variety of tissues, and after high-temperature treatments, its expression was up-regulated in sepals, stamens, pistils, and leaves.
These data show that, overall, the expression levels of most genes of the HDACs were down-regulated after high-temperature treatments, especially in leaves, whereas that of most HAT genes was up-regulated after high-temperature treatment, especially in leaves. In contrast to HDAC genes, the expression of HAT genes RcHAG1, RcHAG2, and RcHAF2 increased after high-temperature treatments in stamens, pistils, and leaves ( Figure 6b, Table S8). Similarly, the expression of RcHAC4 was also up-regulated in petals, stamens, and leaves after high-temperature treatments ( Figure 6; Table S8). RcHAG4 was expressed in a variety of tissues, and after high-temperature treatments, its expression was up-regulated in sepals, stamens, pistils, and leaves.
These data show that, overall, the expression levels of most genes of the HDACs were down-regulated after high-temperature treatments, especially in leaves, whereas that of most HAT genes was up-regulated after high-temperature treatment, especially in leaves.

The Expression Specificity of HDACs and HATs
With the decoding of more and more plants genomes, HDAC and HAT genes families were identified in many plants, such as in Arabidopsis [8], in rice [36,37], in tomato [39], etc.
Histone acetylation plays an important role in plant development and stress responses. These are dynamic and reversible processes mediated by HDACs and HATs enzymes. Our data show that few HDAC and HAT genes with FPKM values greater than 15 exhibit expression patterns in different tissues and organs (i.e., stems, leaves, and flower buds), and in different rose cultivars (Figure 7). We focused on genes for which FPKM values were greater than 15 in stamens, in carpels, in green petals (flower bud stage), in petals changing colors, or in fully developed pink petals of 'OB'. exhibit expression patterns in different tissues and organs (i.e., stems, leaves, and flower buds), and in different rose cultivars (Figure 7). We focused on genes for which FPKM values were greater than 15 in stamens, in carpels, in green petals (flower bud stage), in petals changing colors, or in fully developed pink petals of 'OB'. The HDAC genes, RcHDA6-2, RcHDA19, and RcHDT1 were strongly expressed in roots, stems of 'OB', 'Rw', and 'Ry', while RcHDA14-1 was only strongly expressed in the leaves. All members, except RcHDA6-1, of class I of RPD3/HDA1 subfamily were expressed in the three rose cultivars. Previous report showed that HDA6 acts as a repressor for EIN3-mediated transcription and Jasmonate signaling and suggested that HDA6 was involved in root development [40,41]. RcHDA6-2 was strongly expressed in roots of 'OB', suggesting a putative role in root development in roses, thus in agreement with previously reported data in Arabidopsis.
Interestingly, many RPD/HDA1 members, including RcHDA6-1, RcHDA2-6, RcHDA2-7, RcHDA2-8, RcHDA14-2, RcHDA14-3, and RcHDA14-4, showed very low to no expression in all analyzed organs. Either, these genes are not expressed in these organs, or are not expressed at the analyzed developmental stage and/or growth conditions. RcHDA14-1 was the only class IV gene that showed expression in the leaves of the three The HDAC genes, RcHDA6-2, RcHDA19, and RcHDT1 were strongly expressed in roots, stems of 'OB', 'Rw', and 'Ry', while RcHDA14-1 was only strongly expressed in the leaves. All members, except RcHDA6-1, of class I of RPD3/HDA1 subfamily were expressed in the three rose cultivars. Previous report showed that HDA6 acts as a repressor for EIN3-mediated transcription and Jasmonate signaling and suggested that HDA6 was involved in root development [40,41]. RcHDA6-2 was strongly expressed in roots of 'OB', suggesting a putative role in root development in roses, thus in agreement with previously reported data in Arabidopsis.
Interestingly, many RPD/HDA1 members, including RcHDA6-1, RcHDA2-6, RcHDA2-7, RcHDA2-8, RcHDA14-2, RcHDA14-3, and RcHDA14-4, showed very low to no expression in all analyzed organs. Either, these genes are not expressed in these organs, or are not expressed at the analyzed developmental stage and/or growth conditions. RcHDA14-1 was the only class IV gene that showed expression in the leaves of the three rose cultivars 'OB', 'Rw', and 'Ry', although its exact role in the leaves remains to be elucidated, requiring further studies.
Among the two SIR2 analyzed genes, only RcSRT2 was expressed in 'OB' rose petals during flower opening (Figure 4). Recently, Zhang et al. showed that histone deacetylases SRT1 and SRT2 mediate ethylene-induced transcriptional repression by interacting with the protein EIN2 NUCLEAR ASSOCIATED PROTEIN1 (ENAP1) in seedlings of Arabidopsis [42]. It was reported in roses that ethylene influence flower opening by affecting ethylene receptor and/or signaling genes [42,43]. Therefore, it is possible that in the rose, RcSRT1 and RcSRT2 play a role in flower opening by acting in the ethylene pathway. Therefore, it will be interesting to address the link between RcSRT1 and RcSRT2 and ethylene perception and signaling during flower opening.
Members of the HD2 subfamily were expressed in roots, stems, leaves and flowers, with the highest expression level in flower buds and flowers (Figure 4), especially in young petals in closed buds, in petals at the changing colors development stage and in carpels (FB_GP, FB_CP, and Ca, respectively; Figure 4a). These data indicate that HD2 may play important roles in a diversity of functions during flower development. As support to this hypothesis, in Arabidopsis, HDT1, HDT2, and HDT3 were shown to be important for ovule and embryo development [44], in longan, enhanced expression of DlHD2 was associated with delayed fruit senescence [45], and in O. sativa, overexpression of OsHDT1 was shown to promote early flowering by affecting key flowering time genes [46].
In the GNAT subfamily, RcHAG1, RcHAG2, RcHAG3-1, and MYST subfamily gene RcHAG4 showed pleiotropic expression in most of the analyzed organs, while RcHAG3-2 showed low to no expression (Table S8). In A. thaliana, HAG1 was shown to play an important role in flower development. Arabidopsis lines mutant in HAG1 displayed a variety of pleiotropic growth defects including dwarfism, loss of apical dominance and loss of fertility [47], as well as homeotic conversion of petals into stamens and of sepals into filamentous structures and formation of ectopic carpels [48].
The expression of two CBP genes RcHAC1 and RcHAC4 was very low in the 3 rose cultivars. In Arabidopsis, HAC1 was reported to play a regulatory role in flowering time and in development. Plants knockout for AtHAC1 exhibit delayed flowering and partially reduced fertility [49], and AtHAC1 was shown to function synergistically with AtHAC4, AtHAC5, and AtHAC12 [50]. The expression of RcHAC genes also varied depending on the rose cultivar, thus, indicating putative functional differentiation in different cultivars.

The Antagonistic Expression Patterns of HDACs and HATs
HDACs and HATs play an important role in plant response to stresses, such as high temperature and drought. Roses have a great economic importance worldwide. Their economic value is mainly related to the characteristics of the flower. Here, our data show that HDACs and HATs have different expression patterns in the flower and in the leaf of the three analyzed rose cultivars.
High-temperature treatments of the ancient Chinese rose 'CL' showed that there were differences in gene expression trends between treated and untreated plants. The expression of most HDAC genes was down-regulated after high-temperature treatment in both floral organs and leaves, with a strong down-regulation observed in leaves. In contrast to RcHDA6-2, RcHDA6-1 expression was high in pistils and leaves of 'CL' plants, and upon high-temperature treatments its expression was significantly decreased. This difference in expression patterns and response to high temperature of RcHDA6-1 and RcHDA6-2 may be related to the fact that RcHDA6-2 is expressed at very low level in 'CL' plants. In Arabidopsis, AtHDA6 has been shown to be involved in the response to salt stress [51]. Arabidopsis lines mutant for AtHDA6 were hypersensitive to salt stress, indicating that expression of this gene is important for the salt stress response. An opposite tend appears to occur in the rose, where heat stress correlates with the down-regulation of RcHDA6-1. It is possible that this gene has evolved differently for the response to heat stress in different species. A similar behavior was observed for RcHDA2-1 and RcHDA14-2 whose expression was high in leaves and respectively in pistils or in sepals, and their expression was down-regulated upon heat treatments.
Conversely to other HDACs subfamilies, HD2 genes RcHDT1 and RcHDT2 were expressed in all analyzed organs, thus similar to previous report in A. thaliana, where AtHD2C was shown to be constitutively expressed in all organs [52]. Similar to other HDACs, the expression of both HD2 genes was down-regulated after heat treatment, indicating that both genes may be involved in the response to heat stress. In A. thaliana, HD2 subfamily genes have also been shown to be involved in plant development [19,53] and their expression was repressed by abscisic acid and was associated with decreased sensitivity to salt stress and drought [52]. Our data combined with those of the literature show the importance of HD2 genes in the response to various stresses such as heat, drought, salinity, and cold in plants.
Conversely to HDAC genes, the expression of HAT genes was up-regulated after heat treatments. The highest response was observed in leaves, thus similar to HDACs, but in an opposite way. Previous studies have shown that HATs are involved in plant development, especially in flower development [49,50,52]. Interestingly, after high-temperature treatment, the expression of RcHAG1, RcHAG2, and RcHAF2 was significantly up-regulated in stamens and pistils. Previous study have shown in roses that a high-temperature treatment affect meiosis, and, more specifically, gamete ploidy, leading to diplogametes formation [54]. Taken together with our data, it possible that HAT genes RcHAG2, RcHAG2, RcHAG4, and RcHAF2 have a role in gametes formation, although the direct link between their miss expression in response to high temperature and the formation of diplogamete in roses has not yet been established.
HDACs and HATs jointly regulated histone acetylation, and the expression trends of HDACs and HATs showed opposite changes after high-temperature treatment (Figure 8), indicating that the histone acetylation degrees in rose flowers and leaves are enhanced under high-temperature treatment, which may be one of the ways to cope with hightemperature stress.
Conversely to HDAC genes, the expression of HAT genes was up-regulated after heat treatments. The highest response was observed in leaves, thus similar to HDACs, but in an opposite way. Previous studies have shown that HATs are involved in plant development, especially in flower development [49,50,52]. Interestingly, after high-temperature treatment, the expression of RcHAG1, RcHAG2, and RcHAF2 was significantly up-regulated in stamens and pistils. Previous study have shown in roses that a high-temperature treatment affect meiosis, and, more specifically, gamete ploidy, leading to diplogametes formation [54]. Taken together with our data, it possible that HAT genes RcHAG2, RcHAG2, RcHAG4, and RcHAF2 have a role in gametes formation, although the direct link between their miss expression in response to high temperature and the formation of diplogamete in roses has not yet been established.
HDACs and HATs jointly regulated histone acetylation, and the expression trends of HDACs and HATs showed opposite changes after high-temperature treatment (Figure 8), indicating that the histone acetylation degrees in rose flowers and leaves are enhanced under high-temperature treatment, which may be one of the ways to cope with high-temperature stress.  . Expression patterns of the HDAC (blue) and HAT (red) genes in various organs of R. chinensis 'CL' grown in normal or at high-temperature conditions. Boxes to the left with blue downward-pointing arrow contain genes whose expression level was down-regulated in plants growth at high temperature. Boxes to the right with red upward-pointing arrow contain genes whose expression levels were up-regulated in plants growth at high-temperature treatment.
Moreover, since histone acetylation plays an important role in plant growth and development and resistance to various stress, the fact that the expression levels of histone acetylation-related genes families are modified in rose grown in high-temperature environments, may indicate their important roles in response to heat stress. Therefore, the present work paves the way for further studies to help identify the associated regulatory pathways and specific molecular mechanisms.

Conclusions
In this study, 23 HDAC and 10 HAT genes were identified in R. chinensis 'Old Blush'. Their structures, conserved motifs, physicochemical properties, phylogeny and synteny were studied. The tissue-specific expression of the HDAC and HAT genes was analyzed in three rose cultivars (R. chinensis 'Old Blush', R. wichurana and R. hybrida 'Yesterday'). After heat stress, it showed the antagonistic expression patterns of HDACs and HATs in R. chinensis 'Chilong Hanzhu'. This study paved a way for deciphering the functional roles of RcHDAC and RcHAT genes during plant growth as well as addressing the link between histone acetylation and heat stress response in roses.