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Genome-wide identification of the OVATE gene family and revelation of its expression profile and functional role in eight tissues of Rosa roxburghii Tratt

BMC Plant Biology volume 24, Article number: 1068 (2024) Cite this article

Abstract

Background

The OVATE gene family is a new class of transcriptional repressors, which play an important regulatory role in plant growth and development. Many studies have proved that the OVATE gene family can regulate the development of plant tissues and organs and resist stress, but its quantity and functional role in Rosa roxburghii remain unknown.

Results

In this study, 14 OVATE family members were identified by re-annotating the genome of Rosa roxburghii, and these members were unevenly distributed on 6 chromosomes. Evolutionary analysis indicated that these family members were classified into three groups. In their promoter regions, many hormone-related cis-acting elements such as ABA, GA, and MeJA were identified. Segmental duplication is an important driving force for the expansion of the OVATE family in Rosa roxburghii. Transcriptome sequencing and RT-qPCR analysis showed that OVATE gene family had a specific tissue expression pattern in Rosa roxburghii. For instance, the expression level of gene Rr602241 in leaves was more than 4 times that of other tissues. The gene Rr101515 was highly expressed in FR1 and FR4 stages of fruit tree development, and was highly homologous to the gene regulating fruit shape in tomatoes. These results suggest that members of the OVATE gene family may have diverse functions in different tissues. Furthermore, based on the transcriptome data of eight tissues, a transcriptional regulatory co-expression network of different transcription factors and 14 OVATE genes was constructed.

Conclusion

In conclusion, our study provides the expression profiles of the OVATE family and reveals the potential functional roles of different members in the growth and development of Rosa roxburghii Tratt.

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Introduction

Chestnut rose (Rosa roxburghii Tratt.) is a perennial fruit tree of the Rosaceae Rosae genus that originated in the southwestern region of China [1, 2]. As a traditional food with both medicinal and edible uses, the fruit of the chestnut rose is rich in multiple beneficial components for the human body such as polyphenols, flavonoids, superoxide dismutase, amino acids, and vitamins. Research indicates that the content of ascorbic acid in the fruit of the chestnut rose is more than 2000 mg/100 g of fresh weight [3]. Abundant beneficial substances endow the chestnut rose with many functions such as cancer prevention, immune enhancement, anti-atherosclerosis, and antibacterial. Therefore, its cultivation and consumption have gradually expanded and it has become one of the important economic crops in the southwestern region [4].

Due to the lack of available reference genome sequences, previous studies have focused on the study of functional components in the fruit of Rosa roxburghii Tratt. For example, Lu et al. first investigated the genome of Rosa roxburghii Tratt and identified 11 candidate genes involved in ascorbic acid metabolism, and 17 amino acids were identified in fruits [5, 6]; Based on transcriptome and qPCR analysis, Su et al. [3] identified 13 transcription factors associated with sugar and organic acid metabolism, including sucrose synthase gene RrSUS3; the content of L-Ascorbic acid reached the highest level when the fruit was nearing maturity, while the content in leaves and other tissues was quite low, and the expression level of dehydroascorbate reductase was strongly correlated with the concentration of L-Ascorbic acid [1]. At present, there are relatively few related studies on the molecular genetics of Rosa roxburghii. Based on transcriptome sequencing and De novo assembly technology, Huang et al. [7] discovered 163 Mybs-related genes and 37,545 microsatellite loci in Rosa roxburghii Tratt; Qin et al. [8] identified 38 members of HD-Zip gene family in the ‘Guinong 5’ Rosa roxburghii, and these genes were divided into 4 subfamilies and proved to be involved in regulating the development of trichomes. In 2023, Zong et al. [9] published the first chromosome-level genome of Rosa roxburghii Tratt and identified the two key genes GGP and GalLDH in the ascorbic acid biosynthetic pathway. This provides a foundation for further research on the gene function regulation and molecular breeding of Rosa roxburghii Tratt.

The OVATE family proteins (OFPs) are plant-specific transcription factors that have important functions such as regulating the development of tissues and organs and resisting stress [10, 11]. For example, Guan and Zhou et al. [12, 13] found that a structural variation of about 1.7 Mb downstream of the OVATE family member was closely related to the fruit shape of the peach tree; the OsOFP19 modulates plant architecture by integrating the cell division pattern and brassinosteroid signaling and AtOFP1 and AtOFP4 can combine with a multi-protein transcriptional regulatory complex containing BLH6 and KNAT7 proteins to regulate the formation of secondary cell walls [14, 15]. Protein-protein interaction analysis confirmed that OFPs have close functional connections with many other basic regulatory factors of plant development. Hackbusch et al.‘s [16] research shows that the interaction between AtOFP1 and BLH1 (BLH1 is a member of the TALE family protein) leads to the relocation of BLH1 from the nucleus to the cytoplasmic; overexpression of OsOFP2 was found that it can change the leaf and seed shapes of rice and inhibit the GA level by down-regulating the expression level of OsGA20ox7 [17, 18]; while the OVATE genes in tea trees and peppers may probably participate in the regulation of leaf area and pepper morphology [19, 20]. In addition, studies in rice have shown that members of the OVATE gene family are able to participate in the resistance of plants to cold stress [21]. Fruits are the main harvested products of Rosa roxburghii Tratt trees, and their shapes have important impacts on the yield of the fruit trees. However, as an important regulatory factor for fruit development, the number and function of the OVATE gene family in Rosa roxburghii Tratt have still not been revealed.

In this study, we re-annotated the Rosa roxburghii Tratt genome and thereby conducted comprehensive identification, gene structure, conserved motifs, cis-acting elements, phylogenetic and evolutionary analysis of the members of the OVATE gene family in the genome. Meanwhile, transcriptome sequencing and RT-qPCR analysis of leaf, flower and fruit samples at four different developmental stages were carried out to reveal the expression characteristics of the OVATE gene family in Rosa roxburghii Tratt. In conclusion, this study reveals the evolutionary characteristics and expression profiles in different tissues of the Rosa roxburghii OVATE gene family, which provides important insights for in-depth study of the regulatory mechanism of the OVATE gene family and the breeding of Rosa roxburghii Tratt.

Materials and methods

Plant materials

Eight different tissue samples of wild Rosa roxburghii Tratt were collected from Renhuai City, Guizhou Province, China (E:106.38、N:27.85). These samples include flowers at two developmental stages, leaves, and 5 kinds of fruits at different developmental stages, and their specific codes are as follows: FL1 (Flower bud), FL2 (Flower), LF (Leaf), FR1(the first sampling of the fruit at the growth stage), FR2 (the second sampling of the fruit at the growth stage), FR3 (the third sampling of the fruit at the growth stage), FR4 (the fourth sampling of the fruit at the growth stage), FR5 (the fifth collection of fruits approaching maturity). After these samples were collected, they were immediately transported to the laboratory with dry ice and stored at -80° C before use. All samples contained three biological replicates. The Rosa roxburghii Tratt plant samples used in this study were identified by the local lecturer Qin-Xiao Zhang. A dried plant specimen was kept as a backup at Moutai Institute.

Genome annotation and identification of the OVATE gene family

First, obtain the reference genome and transcriptome sequences of Rosa roxburghii Tratt from the CNGB Nucleotide Sequence Archive database (https://db.cngb.org/cnsa/, accession CNP0004212). Due to the lack of available protein sequences and gff files, the coding protein genes of Rosa roxburghii Tratt were re-predicted and annotated based on the above data. Detailed methods refer to the research of Zong et al. [9]. In order to identify the members of the Rosa roxburghii Tratt OVATE gene family, the Hidden Markov Model (HMM) profile of the OVATE family (PF04844) was downloaded from the InterPro database (https://www.ebi.ac.uk/interpro/download/Pfam/). Subsequently, the hmmsearch (v3.3.2) software and the aforementioned model were used to identify the Rosa roxburghii Tratt OVATE genes [22]. Candidate sequences with e-value < 1e-10 were retained and further verified through SMART online software (https://smart.embl.de/) [23, 24]. The prediction of the genome-wide transcription factors of Rosa roxburghii Tratt was conducted using PlantTFDB v5.0 (https://planttfdb.gao-lab.org/) [25].

Chromosome distribution and sequence analysis

Based on information such as gff files and chromosome length, the chromosomal distribution of all OVATE gene family members is visualized by MG2C website (http://mg2c.iask.in/mg2c_v2.1/) [26]. The molecular weight (MW), number of amino acids and isoelectric point (pI) of all OVATE proteins were predicted using the ExPasy web site (https://web.expasy.org/). Analysis of protein conserved motifs and cis-acting elements were accomplished respectively using TBtools and the PlantCARE online tool (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/) [27, 28]. Finally, the gene structure, motifs and cis-acting elements were visualized by the TBtools software [29, 30].

Phylogenetic tree construction, gene duplication and synteny analysis

The protein sequences of Arabidopsis thaliana (https://www.arabidopsis.org/), peach (Peach Genome Database, http://www.stylebio.cn/index.html) and apple (https://iris.angers.inra.fr/gddh13/) were downloaded and then used for the identification of the members of the OVATE family, and the identified OVATE family members were used together with the Rosa roxburghii Tratt OVATE gene family to conduct multiple sequence alignments using the ClustalW program built in MEGA11. The phylogenetic tree was constructed using the NJ (Neighbor–Joining) method with the following parameters: bootstrap method = 1,000 replicates, p-distance model, pairwise deletion, Uniform rates. All protein sequences were classified based on clustering results. Meantime, the genomes of cotton (https://ngdc.cncb.ac.cn/gwh, under accession no. GWHBISS00000000) and cabbage (BRAD, http://39.100.233.196:82/download_genome/Brassica_Genome_data/Brara_Chiifu_V4.0/) were also downloaded for synteny analysis [31, 32]. Both gene duplication events and synteny analysis (Rosa roxburghii Tratt vs. Arabidopsis thaliana; Rosa roxburghii Tratt vs. Malus domestica Borkh; Rosa roxburghii Tratt vs. Prunus persica; Rosa roxburghii Tratt vs. B. rapa Chiifu v4.0; Rosa roxburghii Tratt vs. Gossypium raimondii) were performed using the default parameters of TBtools software [30, 33].

Transcriptome sequencing and data analysis of eight Rosa roxburghii Tratt tissues

Using the Total RNA Purification Kit (cat DP441, Tiangen, China), the total RNA of all samples was extracted according to the manufacturer’s protocol. Subsequently, agarose gel electrophoresis and the Nanodrop 2500 (Thermo Fisher Scientific, US) instrument were used to detect the quality of RNA, and qualified RNA samples were used for transcriptome sequencing. In order to construct the RNA library, mRNA was enriched with magnetic beads with Oligo (dT), and after randomly fragmenting the mRNA by adding Fragmentation Buffer, the first cDNA strand was synthesized with random hexamers as primers, and then buffer, dNTPs, RNase H and DNA polymerase I were added to synthesize the second cDNA strand. Finally, the cDNA library was obtained through PCR enrichment [34, 35]. The HiSeq 2500 System was used for transcriptome sequencing, and the sequencing read length was PE150.

Before conducting the transcriptome analysis, the raw sequencing data was first filtered using the fastp software [36]. The specific filtering parameters are as follows: --length_required = 25 --cut_front --cut_window_size 4 --cut_mean_quality 15 --compression = 6. After constructing the genomic index using hisat2, the paired-end sequencing data were aligned to the reference genome under default parameters [37]. The sam files were converted to bam and sorted by the santools software [38]. Finally, the StringTie software was used to calculate the FPKM expression level of genes, and the R package DEGseq was used for the analysis of differentially expressed genes (fold_change > 2 and the q-value < 0.05) [39].

The quantitative real-time PCR (RT-qPCR)

RT-qPCR was performed to verify the expression levels of the OVATE gene family members in different tissues of Rosa roxburghii Tratt (the verified genes need to meet the condition that the FPKM value in at least any four tissues was greater than 1). The β-actin gene was used as an internal reference gene [7]. Each sample had three biological replicates [33]. Detailed primer information is listed in Table S1.

Statistical analysis

One-way ANOVA analysis of column chart data was performed using SPSS 22.0 and plotted using Python 3.6. KEGG enrichment analysis and Pearson correlation analysis of OVATE genes and different transcription factors were performed by the omicshare online analysis platform (https://www.omicshare.com/). In this study, there was a statistically significant difference in the use of different letters (P < 0.05). The gene correlation expression networks were constructed using P < 0.05 as a screening parameter and visualized using Cytoscape v3.10 software.

Results

Gene model prediction and annotation

Although the genome sequence at the chromosomal level of Rosa roxburghii Tratt has been published, there is a lack of corresponding proteins and CDS sequences. Therefore, we carried out new prediction and functional annotation for the Rosa roxburghii Tratt gene model. The results show that a total of 40,020 protein-coding genes were predicted, and their detailed protein sequences and CDS sequences are respectively listed in Table S2 and Table S3. Meanwhile, annotation information such as KEGG, GO and UniPort is shown in Table S4. The above results are highly consistent with the genome-reported results, which not only proves the reliability of the annotation results, but also provides a solid foundation for the identification of gene families and transcriptome analysis [9].

Identification and characterization of OVATE gene family in Rosa roxburghii Tratt

In the genome of Rosa roxburghii Tratt, a total of 14 members of the OVATE gene family were identified, and these members were unevenly distributed on 6 chromosomes (Fig. 1). Among them, chromosome 1 and chromosome 6 both contain 3 OVATE genes, chromosome 2 and chromosome 3 contain 2 and 4 OVATE genes respectively, while chromosome 4 and chromosome 7 both contain one OVATE gene. No OVATE gene family members were identified on chromosome 5. Further physicochemical property analysis shows that the number of amino acids of the OVATE gene family is between 104 and 399, and the corresponding molecular weight (MW) varies between 12.23 and 45.42 kDa. The isoelectric point (pI) of these members varies from 4.00 to 9.96. Interestingly, except that Rr602242 is located in the cell wall, all other family members are located in the Nucleus. This indicates that Rr602242 may have undergone functional differentiation from other members (Supplementary Table 5, Table 6).

Fig. 1
figure 1

Distribution of the OVATE genes in the Rosa roxburghii genome. The left axis shows the length of each chromosome, and it was estimated in mega base (Mb)

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Phylogenetic analysis of the OVATE family

In order to reveal the evolutionary relationship of the OVATE gene family in different species, the OVATE protein sequences of Arabidopsis thaliana, apple and peach were collected and identified, and multiple sequence alignments were performed together with the Rosa roxburghii Tratt OVATE protein. The NJ phylogenetic tree shows that all sequences are divided into three groups: Group I, Group II and Group III (Fig. 2). In Group I, the numbers of OVATE family members in Rosa roxburghii Tratt, apple, peach tree and Arabidopsis thaliana are 4, 10, 5 and 7 respectively; while in Group II, the corresponding numbers are 5, 7, 5 and 5 respectively. Meanwhile, in Group III, it respectively contains 5 Rosa roxburghii Tratt, 8 apple, 5 peach tree and 6 Arabidopsis thaliana OVATE family members. These results suggest that the number of OVATE included in Group II is relatively conservative among different species, while the first and third types of OVATE genes in apple may have undergone a unique expansion process.

Fig. 2
figure 2

Construction and analysis of the phylogenetic tree. An unrooted neighbor-joining phylogenetic tree was constructed based on the OVATE sequences in the genomes of Rosa roxburghii Tratt, Arabidopsis thaliana, apple and peach. The bootstrap test was set to 1,000 replicates

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Gene structure, conserved motifs, and cis-acting element analysis

Next, the gene structure, conserved motifs and cis-acting elements are analyzed to reveal the intrinsic relationship of the OVATE gene family. The result is shown as in Fig. 3. Among the members of the Rosa roxburghii Tratt OVATE family, a total of 6 conserved motifs were identified. Among them, motif 1 and motif 2 are highly conserved among all members, while motif 6 only exists in Rr303869 and Rr101515; motif 3 and motif 5 only exist in Rr601085 and Rr306105; and only Rr402601 and Rr203675 contain motif 4 (Fig. 3A). The sequence of 2000 bp upstream of the promoter was extracted for the identification and analysis of cis-acting elements to further understand the regulatory network of the Rosa roxburghii Tratt OVATE gene. In addition to a large number of common light-responsive elements, many cis-acting elements involved in the responses of hormones such as GA (including GARE-motif, P-box and TATC-box), ABA (ABRE element), MeJA (including CGTCA-motif and TGACG-motif) and SA (TCA-element) were identified (Fig. 3B). In addition, in the promoter regions of some family members, cis-acting elements related to responses to stresses (including low-temperature responsive element, TC-rich repeats and WUN-motif) and cell differentiation were also identified. Interestingly, most family members only contain one exon, which is different from the previous research in other species (Fig. 3C) [33].

Fig. 3
figure 3

Motif, gene structure and cis-acting regulatory elements analysis of the OVATE gene family. (A) Protein motifs of OVATE genes. Six conservative motifs are shown in the figure. (B) Cis-acting regulatory elements analysis of the OVATE genes. Sequences of the 2000 bp above the start codon were used to identify cis-acting elements. (C) Gene structure of OVATE genes

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Synteny analysis of OVATE gene family

Gene duplication is one of the important driving forces for the expansion of gene families in the process of plant evolution. Among the members of the OVATE gene family, a total of four duplication events were identified, which are respectively: Rr203675 and Rr402601, Rr204581 and Rr101515, Rr306105 and Rr601085, Rr303869 and Rr602242. Considering the number of the OVATE family, segmental duplication plays an important role in the expansion of the OVATE family (Fig. 4).

Fig. 4
figure 4

Circle plot showing collinearity of OVATE gene family in Rosa roxburghii Tratt. Collinearity genes were highlighted with red curved lines

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In order to further study the evolutionary relationship of the ovate gene family in different plants, we selected five representative plant genomes of Arabidopsis thaliana, peach tree, apple, chinese cabbage and cotton for synteny analysis with Rosa roxburghii. The results showed that 16 (Rrox vs. Arabidopsis), 22 (Rrox vs. peach), 33 (Rrox vs. apple), 12 (Rrox vs. cabbage) and 31 (Rrox vs. cotton) synteny gene pairs were identified respectively (Fig. 5, Supplementary Table S7). In Rosaceae plants, the OVATE gene family in different species may have experienced unique evolutionary processes. Compared with Rosa roxburghii, the members of the OVATE family in apple and cotton have undergone obvious expansion. Some Rosa roxburghii OVATE genes have multiple collinear relationships in apple and cotton, which suggests that the expansion of the OVATE gene family in apple and cotton may have occurred after the differentiation from Rosa roxburghii.

Fig. 5
figure 5

Synteny analysis of OVATE genes from Arabidopsis, Peach and Apple, Cabbage, Cotton

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Transcriptome sequencing and overview of differentially expressed genes

At present, there is a lack of sufficient transcriptome data of different tissues of Rosa roxburghii Tratt being published. In this study, we have carried out transcriptome sequencing for 24 samples from 8 different Rosa roxburghii Tratt tissues. After filtering the raw sequencing data, a total of 168.41 Gb of clean data was obtained, and the clean data of each sample reached 5.88 Gb, and the percentage of Q30 bases was 92.78% or more (Supplementary Table S8). Transcriptome analysis shows that there are a large number of differentially expressed genes among different tissues, and the number is between 1,680 (FR3vFR5) and 9,424 (FR2vFR5) (Fig. 6A). These differentially expressed genes may have promoted the growth, development and metabolite accumulation of different tissues. In order to further classify these differential genes, KEGG enrichment analysis was implemented. The results showed that there were 12,686 differential genes in fruits at different developmental stages, which were mainly enriched in secondary metabolism, hormone signaling, sugar, amino acids, vitamins and MAPK signaling pathways (Fig. 6B); While between the fruits and the flowers, in addition to a large number of pathways such as metabolites, plant hormones and photosynthesis being enriched, pathways such as carotenoid biosynthesis are also enriched. These enriched genes may be involved in regulating the accumulation of carotenoids during the fruit ripening process (Fig. 6C). All the differentially expressed genes are detailedly listed in Supplementary Table S9.

Fig. 6
figure 6

Differentially expressed genes among eight tissues of Rosa roxburghii and KEGG enrichment analysis. (A) Number of differentially expressed genes in different tissues. (B) KEGG enrichment analysis of differentially expressed genes in fruits at five developmental stages. (C) KEGG enrichment analysis of differentially expressed genes between fruits and flowers

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Transcriptome analysis and RT-qPCR verification of OVATE family expression level

Based on the above transcriptome analysis results, we further focused on the expression characteristics of the OVATE gene family in Rosa roxburghii Tratt. The results indicate that the members of the OVATE family in Rosa roxburghii Tratt show diverse expression characteristics (Fig. 7 and Supplementary Figure S1). Among them, Rr100508, Rr101515, Rr203675, Rr303869, Rr601085, Rr602241 and Rr204581 have relatively high expression levels in the first (FL1) or second stage (FL2) of flower development; while Rr300983, Rr602241 and Rr602242 are highly expressed in leaves, especially the latter whose expression level in leaves is significantly higher than that in other tissues, suggesting that this gene is closely related to leaf development. During the five stages of fruit development, the expression levels of the members of the OVATE family are different. For instance, Rr100508 and Rr101515 have higher expression levels in FR1 and FR2, Rr601085 is highly expressed in FR3, FR4 and FR5, while Rr203675 shows a gradually increasing trend along with the development of the fruit. Especially Rr306105, the expression level increased sharply during the rapid expansion period of the fruit (FR3), which means it might be a key gene regulating the size of Rosa roxburghii Tratt fruit.

Fig. 7
figure 7

The expression characteristics of OVATE gene family in eight tissues of Rosa roxburghii were analyzed based on transcriptome sequencing. Different letters above the bars represent significant differences at p < 0.05. ND means not detected

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Above, we analyzed the expression levels of the OVATE gene family in Rosa roxburghii Tratt through transcriptome data. However, the analysis methods, sequencing depth, and experimental design can all affect the accuracy of the analysis results. Therefore, we selected 6 pairs of primers to further verify their true expression levels through RT-qPCR. It should be pointed out that some family members have extremely low expression levels or no expression in Rosa roxburghii Tratt, making it difficult to verify their true expression characteristics through RT-qPCR (Fig. 7 and Supplementary Figure S1). The results are shown in Fig. 8, Rr100508, Rr101515, Rr203675, Rr601085 and Rr602241 have relatively high expression levels at different developmental stages of flowers; while Rr101515, Rr601085 and Rr306105 are highly expressed at a certain stage of fruit development. Overall, these results are highly consistent with the transcriptome data, reflecting the authenticity and reliability of the data.

Fig. 8
figure 8

Expression of OVATE genes analyzed by RT-qPCR in different tissues of Rosa roxburghii. Different letters above the bars represent significant differences at p < 0.05. ND means not detected

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Constructing the transcriptional regulatory network of the OVATE gene family

To further reveal the transcriptional regulatory network of the OVATE gene family members in different tissues of Rosa roxburghii Tratt, we identified 1,592 transcription factors from 56 transcription factor families and analyzed their correlation expressions with the members of the OVATE family. Finally, a regulatory network was constructed based on the significantly associated members (Supplementary Table S10, P < 0.05) (Fig. 9). The results indicate that each OVATE gene has transcription factors that are significantly correlated with its expression, and the categories of these transcription factors are diverse. Among them, a large number of members of MYB, MYB_related, ERF, NAC, bHLH, bZIP, WRKY, FAR1, C2H2, B3, and C3H have significant co-expression relationships with the OVATE family. These results suggest that in different tissues and developmental stages, the OVATE gene family may be regulated by different transcription factors. At the same time, some members of the family whose expression levels are significantly related may have similar regulatory functions (Supplementary Figure S2).

Fig. 9
figure 9

Transcriptional regulatory network of OVATE gene family in Rosa roxburghii. The correlations between the expression of OVATE genes and that of different transcription factors are shown with colored lines (Pearson’s correlation test, P ≤ 0.05). The detailed expression correlations used for network construction are listed in Table S10

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Discussion

The progress of sequencing technology has greatly promoted the progress of genomics, and the genomes of many Rosaceae plants, including Rosa roxburghii Tratt, have been released [40]. Due to the lack of available protein sequences, we re-predicted and annotated the gene model based on the published genome of Rosa roxburghii Tratt, and the highly consistent annotation results provided a basis for accurately identifying the gene family of Rosa roxburghii Tratt and further utilization [9]. The OVATE gene family was first discovered in tomatoes and has important functions in regulating plant organ morphology and resisting various stresses [17]. In recent years, it has attracted the attention of more and more researchers. For example, studies in bananas have found that MaOFP1 can regulate the ripening of banana fruits [41]; Ding et al. [40] analyzed the OVATE gene family in six Rosaceae plants, including pear trees, and the results showed that all OFP genes were divided into eight categories, and the PbrOFP genes may contain specific regulatory mechanisms in tissues such as flowers, ovaries and fruits, and five PbrOFP genes may play a key role in the defense against stress. In this study, we identified a total of 14 OVATE genes from the genome of Rosa roxburghii Tratt, and these genes were unevenly distributed on the six chromosomes of Rosa roxburghii Tratt. Overall, the number of OVATE family members in Rosa roxburghii Tratt was approximately equivalent to that in wild tea trees, but less than that in pear trees [19, 40].

Physicochemical analysis showed that the molecular weight of the OVATE family was between 12.23 and 45.42 kDa, and all other family members were localized in the nucleus except Rr602242, which was predicted to be located in the cell wall. To further reveal the evolutionary characteristics of the OVATE family in different species, we collected the OVATE sequences from the genomes of Arabidopsis thaliana (18), peach trees (15) and apples (25) and constructed a neighbor-joining phylogenetic tree together with Rosa roxburghii Tratt. The results showed that all the OVATE sequences were classified into three group, among which the OVATE sequences of peach tree and Rosa roxburghii Tratt showed high homology, while the OVATE members in apple genome expanded significantly [42]. Meanwhile, four pairs of segmental duplication genes were identified. Considering the number of OVATE genes in Rosa roxburghii Tratt, segmental duplication may play an important role in the evolution of the OVATE family in Rosaceae plants. Interestingly, the types of conserved motifs identified in either Rosa roxburghii Tratt or peach trees are fewer than those in tea plants, sorghum and peppers [19, 20, 43]. In addition, a large number of cis-acting elements related to ABA, GA, Auxin, MeJA, and MYB were identified in the promoter region of the sequence, suggesting that the OVATE gene family may assume a variety of functions.

To understand the expression characteristics of OVATE family members in different tissues and developmental stages of Rosa roxburghii Tratt, transcriptome sequencing of eight organs was performed. Transcriptome data analysis identified a large number of differentially expressed genes among different tissues, and these genes were mainly enriched in related pathways such as secondary metabolites and plant hormone signal transduction. Here, we focus on the expression characteristics of the OVATE family. The results indicated that the members of the OVATE family in Rosa roxburghii Tratt exhibited diverse expression characteristics in eight tissues. For example, the gene Rr106085 was highly expressed in flower (FL2), leaf (LF) and fruit development (FR1, FR2), Rr300983 was highly expressed in leaf and fruit development FR2 stage, and Rr306105 was sharply expressed during rapid fruit expansion (FR3). The aforementioned genes and OFP4 (AT1G06920) were all classified into Group III, while the latter has been confirmed to regulate cell wall formation [15]. Further analysis showed that genes Rr106085 and Rr306105 were highly homologous to SlOFP20, which regulates flower and pollen development in tomatoes [10, 44]. While Rr101515 is highly homologous to the OVATE member that regulates fruit shape in tomato [45]. This strongly indicates that the genes in these Rosa roxburghii Tratt may also have similar functions in regulating organ development. In addition, we verified the expression characteristics of 6 OVATE gene family members by RT-qPCR, and the results were highly consistent with the transcriptome data, demonstrating the high quality of transcriptome sequencing and the reliability of the analysis results.

Previous studies have demonstrated that members of the OVATE family can bind to the KNAT7 protein or exert regulatory effects in synergy with the TALE protein. However, there is still a lack of research on the transcriptional regulatory network of the OVATE gene family [15, 16]. Transcriptional regulatory network analysis showed that the OVATE gene family was correlated with the transcription factor families such as MYB, MYB_related, ERF, NAC, bHLH, and WRKY, which may confer the diversity and specificity of the spatiotemporal expression of OVATE gene. However, the in-depth regulatory mechanism remains to be further revealed.

Conclusions

To identify the OVATE gene family, we first performed gene model prediction and annotation for Rosa roxburghii Tratt. Based on this, a total of 14 OVATE genes were identified on the six chromosomes of Rosa roxburghii Tratt. These genes are divided into three classes, each of which contains conserved motif 1 and motif 2 and contains a large number of hormone-related cis-acting elements in their promoter regions. The segmental duplication plays an important role in the expansion of the OVATE family. Meanwhile, we conducted transcriptome sequencing and analysis on eight organs such as flowers, leaves, and fruits of Rosa roxburghii Tratt, and a large number of differentially expressed genes were identified. Members of the OVATE gene family show significant differences in expression levels in different tissues, and these genes may regulate specific tissue development. The RT-qPCR verification results were highly consistent with the transcriptome data, indicating the reliability of the research results of this study. These results will help reveal the potential biological functions of the OVATE gene family and provide clues for further in-depth verification in the future.

Data availability

The raw sequence data reported in this paper have been deposited in the Genome Sequence Archive (Genomics, Proteomics & Bioinformatics 2021) in National Genomics Data Center (Nucleic Acids Res 2022), China National Center for Bioinformation / Beijing Institute of Genomics, Chinese Academy of Sciences (GSA: CRA017453) that are publicly accessible at https://ngdc.cncb.ac.cn/gsa.

References

  1. Huang M, Xu Q, Deng X-X. l-Ascorbic acid metabolism during fruit development in an ascorbate-rich fruit crop chestnut rose (Rosa roxburghii Tratt). J Plant Physiol. 2014;171(14):1205–16.

    Article  CAS  PubMed  Google Scholar 

  2. Su L, Zhang T, Cheng Z-M. Identification of transcription factors contributing to vitamin C synthesis during Rosa roxburghii fruit development by integrating transcriptomics and metabolomics. Hortic Plant J 2024.

  3. Su L, Zhang T, Wu M, Zhong Y, Cheng Z. Transcriptome and Metabolome Reveal Sugar and Organic Acid Accumulation in Rosa roxburghii Fruit. Plants 2023, 12(17).

  4. Xu L, Yang H, Li C, Liu S, Zhao H, Liao X, Zhao L. Composition analysis of free and bound phenolics in chestnut rose (Rosa roxburghii Tratt.) Fruit by UHPLC-IM-QTOF and UPLC-QQQ. Lwt 2023, 185.

  5. Lu Min AH, Li L. Genome survey sequencing for the characterization of the genetic background of Rosa roxburghii Tratt and Leaf Ascorbate metabolism genes. PLoS ONE 2016, 11(2).

  6. Lu M, An H, Wang D. Characterization of amino acid composition in fruits of three Rosa roxburghii genotypes. Hortic Plant J. 2017;3(6):232–6.

    Article  Google Scholar 

  7. Remington DL, Huang X, Yan H, Zhai L, Yang Z, Yi Y. Characterization of the Rosa roxburghii Tratt transcriptome and analysis of MYB genes. PLoS ONE 2019, 14(3).

  8. Qin J, Nan H, Ma W, Zhang J, Lu J, Wu A, Lu M, An H. Genome wide characterization and identification of candidate HD-Zip genes involved in prickle density in Rosa roxburghii. Sci Hort 2024, 330.

  9. Zong D, Liu H, Gan P, Ma S, Liang H, Yu J, Li P, Jiang T, Sahu SK, Yang Q, et al. Chromosomal-scale genomes of two Rosa species provide insights into genome evolution and ascorbate accumulation. Plant J. 2023;117(4):1264–80.

    Article  PubMed  Google Scholar 

  10. Zhou S, Hu Z, Li F, Tian S, Zhu Z, Li A, Chen G. Overexpression of SlOFP20 affects floral organ and pollen development. Hortic Res 2019, 6(1).

  11. Zsogon A, Cermak T, Naves ER, Notini MM, Edel KH, Weinl S, Freschi L, Voytas DF, Kudla J, Peres LEP. De novo domestication of wild tomato using genome editing. Nat Biotechnol 2018.

  12. Guan J, Xu Y, Yu Y, Fu J, Ren F, Guo J, Zhao J, Jiang Q, Wei J, Xie H. Genome structure variation analyses of peach reveal population dynamics and a 1.67 mb causal inversion for fruit shape. Genome Biol. 2021;22(1):13.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Zhou H, Ma R, Gao L, Zhang J, Zhang A, Zhang X, Ren F, Zhang W, Liao L, Yang Q et al. A 1.7-Mb chromosomal inversion downstream of a PpOFP1 gene is responsible for flat fruit shape in peach. Plant Biotechnol J 2020.

  14. Yang C, Ma Y, He Y, Tian Z, Li J. OsOFP19 modulates plant architecture by integrating the cell division pattern and brassinosteroid signaling. Plant J. 2018;93(3):489–501.

    Article  CAS  PubMed  Google Scholar 

  15. Li E, Wang S, Liu Y, Chen J-G, Douglas CJ. OVATE FAMILY PROTEIN4 (OFP4) interaction with KNAT7 regulates secondary cell wall formation in Arabidopsis thaliana. Plant J. 2011;67(2):328–41.

    Article  CAS  PubMed  Google Scholar 

  16. Uhrig JHKRJMFSJF. A central role of Arabidopsis thaliana ovate family proteins in networking and subcellular localization of 3-aa loop extension homeodomain proteins. PNAS 2004.

  17. Liu D, Sun W, Yuan Y, Zhang N, Hayward A, Liu Y, Wang Y. Phylogenetic analyses provide the first insights into the evolution of OVATE family proteins in land plants. Ann Botany. 2014;113(7):1219–33.

    Article  CAS  Google Scholar 

  18. Schmitz AJ, Begcy K, Sarath G, Walia H. Rice Ovate family protein 2 (OFP2) alters hormonal homeostasis and vasculature development. Plant Sci. 2015;241:177–88.

    Article  CAS  PubMed  Google Scholar 

  19. An Y, Xia X, Jing T, Zhang F. Identification of gene family members and a key structural variation reveal important roles of OVATE genes in regulating tea (Camellia sinensis) leaf development. Front Plant Sci 2022, 13.

  20. Luo Y, Yang S, Luo X, Li J, Li T, Tang X, Liu F, Zou X, Qin C. Genome-wide analysis of OFP gene family in pepper (Capsicum annuum L). Front Genet 2022, 13.

  21. Ma Y, Yang C, He Y, Tian Z, Li J, Sunkar R. Rice OVATE family protein 6 regulates plant development and confers resistance to drought and cold stresses. J Exp Bot. 2017;68(17):4885–98.

    Article  CAS  PubMed  Google Scholar 

  22. Xiong R, Peng Z, Zhou H, Xue G, He A, Yao X, Weng W, Wu W, Ma C, Bai Q et al. Genome-wide identification, structural characterization and gene expression analysis of the WRKY transcription factor family in pea (Pisum sativum L). BMC Plant Biol 2024, 24(1).

  23. Ahmad S, Ahmad N, Bayar J. Genome-wide identification, characterization, and expression analysis of the Ovate family protein in Oryza sativa under biotic and abiotic stresses. Plant Stress 2023, 10.

  24. Rasool F, Uzair M, Naeem MK, Rehman N, Afroz A, Shah H, Khan MR. Phenylalanine Ammonia-Lyase (PAL) Genes Family in Wheat (Triticum aestivum L.): Genome-Wide Characterization and Expression Profiling. Agronomy 2021, 11(12).

  25. Guo AY, Chen X, Gao G, Zhang H, Zhu QH, Liu XC, Zhong YF, Gu X, He K, Luo J. PlantTFDB: a comprehensive plant transcription factor database. Nucleic Acids Res. 2007;36(Database):D966–9.

    Article  PubMed  PubMed Central  Google Scholar 

  26. Zhang F, Jiang S, Li Q, Song Z, Yang Y, Yu S, Nie Z, Chu M, An Y. Identification of the ALMT gene family in the potato (Solanum tuberosum L.) and analysis of the function of StALMT6/10 in response to aluminum toxicity. Front Plant Sci 2023, 14.

  27. Chen C, Chen H, Zhang Y, Thomas HR, Frank MH, He Y, Xia R. TBtools - an integrative toolkit developed for interactive analyses of big biological data. Mol Plant 2020.

  28. Huang D, Mao Y, Guo G, Ni D, Chen L. Genome-wide identification of PME gene family and expression of candidate genes associated with aluminum tolerance in tea plant (Camellia sinensis). BMC Plant Biol 2022, 22(1).

  29. Zhang F, Yuan A, Nie Z, Chu M, An Y. Identification of the potato (Solanum tuberosum L.) P-type ATPase gene family and investigating the role of PHA2 in response to Pep13. Front Plant Sci 2024, 15.

  30. Chen C, Wu Y, Li J, Wang X, Zeng Z, Xu J, Liu Y, Feng J, Chen H, He Y. TBtools-II: a one for all, all for one bioinformatics platform for biological big-data mining. Mol Plant. 2023;16(11):1733–42.

    Article  CAS  PubMed  Google Scholar 

  31. Zhang L, Liang J, Chen H, Zhang Z, Wu J, Wang X. A near-complete genome assembly of Brassica rapa provides new insights into the evolution of centromeres. Plant Biotechnol J. 2023;21(5):1022–32.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Huang G, Bao Z, Feng L, Zhai J, Wendel JF, Cao X, Zhu Y. A telomere-to-telomere cotton genome assembly reveals centromere evolution and a Mutator transposon-linked module regulating embryo development. Nat Genet. 2024;56(9):1953–63.

    Article  CAS  PubMed  Google Scholar 

  33. An Y, Mi X, Xia X, Qiao D, Yu S, Zheng H, Jing T, Zhang F. Genome-wide identification of the PYL gene family of tea plants (Camellia sinensis) revealed its expression profiles under different stress and tissues. BMC Genomics 2023, 24(1).

  34. Cao K, Ding T, Mao D, Zhu G, Fang W, Chen C, Wang X, Wang L. Transcriptome analysis reveals novel genes involved in anthocyanin biosynthesis in the flesh of peach. Plant Physiol Biochem. 2018;123:94–102.

    Article  CAS  PubMed  Google Scholar 

  35. Kim J, Kang SH, Park SG, Yang TJ, Lee Y, Kim OT, Chung O, Lee J, Choi JP, Kwon SJ, et al. Whole-genome, transcriptome, and methylome analyses provide insights into the evolution of platycoside biosynthesis in Platycodon grandiflorus, a medicinal plant. Hortic Res. 2020;7:112.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Chen S, Zhou Y, Chen Y, Gu J. Fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics. 2018;34(17):i884–90.

    Article  PubMed  PubMed Central  Google Scholar 

  37. Pertea M, Kim D, Pertea GM, Leek JT, Salzberg SL. Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown. Nat Protoc. 2016;11(9):1650–67.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R. The sequence Alignment/Map format and SAMtools. Bioinformatics. 2009;25(16):2078–9.

    Article  PubMed  PubMed Central  Google Scholar 

  39. Wang L, Feng Z, Wang X, Wang X, Zhang X. DEGseq: an R package for identifying differentially expressed genes from RNA-seq data. Bioinformatics. 2010;26(1):136–8.

    Article  PubMed  Google Scholar 

  40. Ding B, Hu C, Feng X, Cui T, Liu Y, Li L. Systematic analysis of the OFP genes in six Rosaceae genomes and their roles in stress response in Chinese pear (Pyrus Bretschneideri). Physiol Mol Biology Plants. 2020;26(10):2085–94.

    Article  CAS  Google Scholar 

  41. Xu C, Liu J, Zhang J, Hu W, Miao H, Zhang J, Jia C, Wang Z, Xu B, Jin Z. Banana Ovate family protein MaOFP1 and MADS-Box protein MuMADS1 Antagonistically Regulated Banana Fruit Ripening. PLoS ONE 2015, 10(4).

  42. Li H, Dong Q, Zhu X, Zhao Q, Ran K. Genome-wide identification, expression, and interaction analysis for ovate family proteins in peach. Mol Biol Rep. 2019;46(4):3755–64.

    Article  CAS  PubMed  Google Scholar 

  43. An Y, Xia X, Zhang X, Liu L, Jiang S, Jing T, Zhang F. Genome-wide identification of the sorghum OVATE gene family and revelation of its expression characteristics in sorghum seeds and leaves. 2024.

  44. Huang Z, Van Houten J, Gonzalez G, Xiao H, van der Knaap E. Genome-wide identification, phylogeny and expression analysis of SUN, OFP and YABBY gene family in tomato. Mol Genet Genomics. 2013;288(3–4):111–29.

    Article  CAS  PubMed  Google Scholar 

  45. Tanksley JLJVEBCaSD. A new class of regulatory genes underlying the cause of pear-shaped tomato fruit. PNAS; 2002.

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Funding

This research was funded by the Natural Science Foundation of Guizhou Province (ZK[2023]453), research Foundation for Scientific Scholars of Moutai Institute (mygccrc [2022]074, mygccrc [2022]077, mygccrc [2022]088, mygccrc[2022]089), the National Natural Science Foundation of China (32160441).

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Author notes

  1. Yanlin An and Xueqi Li contributed equally to this work.

Authors and Affiliations

  1. Department of Food Science and Engineering, Moutai Institute, Luban Street, Renhuai, Guizhou, 564502, P.R. China

    Yanlin An, Xueqi Li, Yani Chen, Sixia Jiang & Feng Zhang

  2. State Key Laboratory of Tea Plant Biology and Utilization, Anhui Agricultural University, 130 Changjiang West Road, Hefei, China

    Tingting Jing

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  2. Xueqi Li
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Contributions

Feng Zhang and Tingting Jing: Conceived and designed the research project. Yanlin An and Xueqi Li: Writing original draft, sample collection, and DNA and RNA extraction, visualization and data curation. Yani Chen and Sixia Jiang: Gene family identification, transcriptome analysis, and RT-qPCR validation. All authors have read and agreed to the published version of the manuscript.

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Correspondence to Tingting Jing or Feng Zhang.

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Not applicable. The sampling of plant material was performed in compliance with institutional guidelines. The research conducted in this study required neither approval from an ethics committee, nor involved any human or animal subjects.

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12870_2024_5775_MOESM1_ESM.xlsx

Supplementary Material 1: Table S1. Primer sequence used in RT-qPCR. Table S2. Whole genome protein sequence of Rosa roxburghii Tratt. Table S3 The CDS sequences of the whole genome of Rosa roxburghii Tratt. Table S4 Gene annotation information of the whole genome of Rosa roxburghii Tratt. Table S5 Sequence characteristics of OVATE genes. Table S6. The cds/protein sequences of the OVATE gene family in Rosa roxburghii. Table S7 Collinear gene pairs among different genomes. Table S8 The data volume and quality value of clean reads in transcriptome sequencing. Table S9 Detailed list of differential genes among different tissues. Table S10 Expression correlation of OVATE members used for co-expression network construction with different transcription factor genes.

12870_2024_5775_MOESM2_ESM.pdf

Supplementary Material 2: Figure S1 The expression characteristics of OVATE gene family in eight tissues of Rosa roxburghii Tratt were analyzed based on transcriptome sequencing. Different letters above the bars represent significant differences at p <0.05. Figure S2 Expression correlation among members of the OVATE gene family based on transcriptome analysis (Pearson’s correlation test, P<0.05)

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An, Y., Li, X., Chen, Y. et al. Genome-wide identification of the OVATE gene family and revelation of its expression profile and functional role in eight tissues of Rosa roxburghii Tratt. BMC Plant Biol 24, 1068 (2024). https://doi.org/10.1186/s12870-024-05775-1

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Keywords

BMC Plant Biology

ISSN: 1471-2229

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