Comparative transcriptome analysis reveals the impact of daily temperature difference on male sterility in photo-thermo-sensitive male sterile wheat.

BACKGROUND: Photo-thermo-sensitive male sterility (PTMS), which refers to the male sterility triggered by variations in photoperiod and temperature, is a crucial element in the wheat two-line hybrid system. The development of safe production and efficient propagation for male sterile lines holds utmost importance in two-line hybrid wheat. Under the stable photoperiod condition, PTMS is mainly induced by high or low temperatures in wheat, but the effect of daily temperature difference (DTD) on the fertility conversion of PTMS lines has not been reported. Here, three BS type PTMS lines including BS108, BS138, and BS366, as well as a control wheat variety J411 were used to analyze the correlation between fertility and DTD using differentially sowing tests, photo-thermo-control experiments, and transcriptome sequencing. RESULTS: The differentially sowing tests suggested that the optimal sowing time for safe seed production of the three PTMS lines was from October 5th to 25th in Dengzhou, China. Under the condition of 12 h 12 °C, the PTMS lines were greatly affected by DTD and exhibited complete male sterility at a temperature difference of 15 °C. Furthermore, under different temperature difference conditions, a total of 20,677 differentially expressed genes (DEGs) were obtained using RNA sequencing. Moreover, through weighted gene co-expression network analysis (WGCNA) and KEGG enrichment analysis, the identified DEGs had a close association with "starch and sucrose metabolism", "phenylpropanoid biosynthesis", "MAPK signaling pathway-plant", "flavonoid biosynthesis", and "cutin, and suberine and wax biosynthesis". qRT-PCR analysis showed the expression levels of core genes related to KEGG pathways significantly decreased at a temperature difference of 15 ° C. Finally, we constructed a transcriptome mediated network of temperature difference affecting male sterility. CONCLUSIONS: The findings provide important theoretical insights into the correlation between temperature difference and male sterility, providing guidance for the identification and selection of more secure and effective PTMS lines.

Microspore-expressed SCULP1 is required for p-coumaroylation of sporopollenin, exine integrity, and pollen development in wheat.

Sporopollenin is one of the most structurally sophisticated and chemically recalcitrant biopolymers. In higher plants, sporopollenin is the dominant component of exine, the outer wall of pollen grains, and contains covalently linked phenolics that protect the male gametes from harsh environments. Although much has been learned about the biosynthesis of sporopollenin precursors in the tapetum, the nutritive cell layer surrounding developing microspores, little is known about how the biopolymer is assembled on the microspore surface. We identified SCULP1 (SKS clade universal in pollen) as a seed plant conserved clade of the multicopper oxidase family. We showed that SCULP1 in common wheat (Triticum aestivum) is specifically expressed in the microspore when sporopollenin assembly takes place, localized to the developing exine, and binds p-coumaric acid in vitro. Through genetic, biochemical, and 3D reconstruction analyses, we demonstrated that SCULP1 is required for p-coumaroylation of sporopollenin, exine integrity, and pollen viability. Moreover, we found that SCULP1 accumulation is compromised in thermosensitive genic male sterile wheat lines and its expression partially restored exine integrity and male fertility. These findings identified a key microspore protein in autonomous sporopollenin polymer assembly, thereby laying the foundation for elucidating and engineering sporopollenin biosynthesis.

Identification and validation of major-effect quantitative trait locus QMS-5B associated with male sterility in photo-thermo-sensitive genic male sterile wheat.

KEY MESSAGE: QMS-5B, a major QTL for photo-thermo-sensitive genic male sterility in wheat, was fine mapped in a 2.15 Mb region harboring a serine/threonine protein kinase gene TraesCS5B03G0887500, which was the most likely candidate gene. Genic male sterility is an essential trait in the utilization of heterosis and hybrid seed production for wheat. Currently, genic male sterile genes have been reported in wheat mutants, but the sterile genes controlling photo-thermo-sensitive genic male sterility in wheat have not been studied systematically. Here, 235 doubled haploid lines derived from a cross between photo-thermo-sensitive genic male sterile line BS462 and its restorer line CP279 were used to map male sterile gene by GenoBaits® Wheat 100 K Panel, bulked segregant exome sequencing (BSE-Seq) and wheat 660 K array. As a result, the major stable QTL on chromosome 5B, QMS-5B, was identified in all four environments, accounting for 7.3-36.4% of the phenotypic variances. Ulteriorly, QMS-5B was delimited to an approximate 2.15 Mb physical interval between KASP-5B5 and KASP-5B6 using kompetitive allele-specific PCR (KASP) markers. Within the interval, twenty-nine high-confidence genes were predicted according to Chinese Spring RefSeq v2.1. TraesCS5B03G0887500, encoding a serine/threonine protein kinase, was identified as the most likely candidate gene for QMS-5B based on weighted gene co-expression network analysis. Expression analysis confirmed that TraesCS5B03G0887500 was significantly differentially expressed in anthers of BS462 and CP279 at different stages under fertile and sterile environments. In addition, flanking KASP marker KASP-5B6 can effectively genotype male sterile lines and restorer lines, and can be used for molecular marker-assisted selection. This study provides insights into for exploring the mechanism of photo-thermo-sensitive genic male sterility in wheat.

Efficacy of Femoral Neck System Fixation vs KHS Femoral Neck Dynamic Compression Locking Plate System in Femoral Neck Fracture.

Background: Femoral neck fracture is acknowledged as one of the common injuries in clinical orthopedics. Our study was aimed at investigating the efficacy of femoral neck fixation vs the KHS dynamic compression locking plate system in the treatment of femoral neck fracture. Methods: This was a prospective study. A toteal of 90 patients with femoral neck fracture who were admitted to The Third Hospital of Hebei Medical University in Shijiazhuang, China from August 2017 to March 2020 were enrolled in our study. The patients were randomly divided into the control group (45 patients allocated to intervention with the novel femoral neck dynamic compression locking plate system) and the study group (45 patients who underwent femoral neck system fixation). Intraoperative blood loss, surgery duration, fracture healing time and related complications in the 2 groups were monitored and evaluated. The recovery of hip joint function at different times in the 2 groups were closely monitored. Results: The 2 groups completed the surgery process, and the incision healed. All patients were followed up for 6 to 8 months, with an average follow-up time of 7.01 ± 0.21 months. Surgery duration, length of hospital stay and fracture healing time in the study group were significantly lower than in the control group (P < .05), while no significant difference was found in intraoperative blood loss between the 2 groups (P > .05). At 1 and 3 months after surgery, hip joint function in the study group was significantly higher than in the control group (P < .05), but 6 months after surgery, there was no significant difference between the 2 groups (P > .05). There were no complications in the study group, whereas 1 patient had a complication in the control group. The total incidence of complications in the study group was lower than in the control group, but the difference was not significant (P > .05). Conclusion: Femoral neck system fixation demonstrated superior efficacy to the KHS femoral neck dynamic compression locking plate system in femoral neck fracture, and is considered as a valid method for wide application.

Comprehensive molecular evaluation of the histone methyltransferase gene family and their important roles in two-line hybrid wheat.

BACKGROUND: Histone methylation usually plays important roles in plant development through post-translational regulation and may provide a new visual field for heterosis. The histone methyltransferase gene family has been identified in various plants, but its members and functions in hybrid wheat related in heterosis is poorly studied. RESULTS: In this study, 175 histone methyltransferase (HMT) genes were identified in wheat, including 152 histone lysine methyltransferase (HKMT) genes and 23 protein arginine methyltransferase (PRMT) genes. Gene structure analysis, physicochemical properties and subcellular localization predictions of the proteins, exhibited the adequate complexity of this gene family. As an allohexaploid species, the number of the genes (seven HKMTs orthologous groups and four PRMTs orthologous groups) in wheat were about three times than those in diploids and showed certain degrees of conservation, while only a small number of subfamilies such as ASH-like and Su-(var) subfamilies have expanded their members. Transcriptome analysis showed that HMT genes were mainly expressed in the reproductive organs. Expression analysis showed that some TaHMT genes with different trends in various hybrid combinations may be regulated by lncRNAs with similar expression trends. Pearson correlation analysis of the expression of TaHMT genes and two yield traits indicated that four DEGs may participate in the yield heterosis of two-line hybrid wheat. ChIP-qPCR results showed that the histone modifications (H3K4me3, H3K36me3 and H3K9ac) enriched in promoter regions of three TaCCA1 genes which are homologous to Arabidopsis heterosis-related CCA1/LHY genes. The higher expression levels of TaCCA1 in F1 than its parents are positive with these histone modifications. These results showed that histone modifications may play important roles in wheat heterosis. CONCLUSIONS: Our study identified characteristics of the histone methyltransferase gene family and enhances the understanding of the evolution and function of these members in allohexaploid wheat. The causes of heterosis of two-line hybrid wheat were partially explained from the perspective of histone modifications.

Global survey of alternative splicing and gene modules associated with fertility regulation in a thermosensitive genic male sterile wheat.

Thermosensitive genic male sterile (TGMS) wheat lines are the core of two-line hybrid systems. Understanding the mechanism that regulates male sterility in TGMS wheat lines is helpful for promoting wheat breeding. Several studies have obtained information regarding the mechanisms associated with male sterility at the transcriptional level, but it is not clear how the post-transcriptional process of alternative splicing might contribute to controlling male sterility. In this study, we performed genome-wide analyses of alternative splicing during the meiosis stage in TGMS line BS366 using PacBio and RNA-Seq hybrid sequencing. Cytological observations indicated that cytoskeleton assembly in pollen cells, calcium deposition in pollen and tapetal cells, and vesicle transport in tapetal cells were deficient in BS366. According to our cytological findings, 49 differentially spliced genes were isolated. Moreover, 25 long non-coding RNA targets and three bHLH transcription factors were identified. Weighted gene co-expression network analysis detected four candidate differentially spliced genes that had strong co-relation with the seed setting percentage, which is the direct representation of male sterility in BS366. In this study, we obtained comprehensive data regarding the alternative splicing-mediated regulation of male sterility in TGMS wheat. The candidates identified may provide the molecular basis for an improved understanding of male sterility.

Comparative Transcriptome Analysis Reveals Key Insights into Fertility Conversion in the Thermo-Sensitive Cytoplasmic Male Sterile Wheat.

Thermo-sensitive cytoplasmic male sterility (TCMS) plays a crucial role in hybrid production and hybrid breeding; however, there are few studies on molecular mechanisms related to anther abortion in the wheat TCMS line. In this study, FA99, a new wheat thermo-sensitive cytoplasmic male sterility line, was investigated. Fertility conversion analysis showed that FA99 was mainly controlled by temperature, and the temperature-sensitive stage was pollen mother cell formation to a uninucleate stage. Further phenotypic identification and paraffin section showed that FA99 was characterized by indehiscent anthers and aborted pollen in a sterile environment and tapetum was degraded prematurely during the tetrad period, which was the critical abortion period of FA99. The contents of O2-, H2O2, MDA and POD were significantly changed in FA99 under a sterile environment by the determination of physiological indexes. Furthermore, through transcriptome analysis, 252 differentially expressed genes were identified, including 218 downregulated and 34 upregulated genes. Based on KOG function classification, GO enrichment and KEGG pathways analysis, it was evident that significant transcriptomic changes in FA99 under different fertility environments, and the major differences were "phenylalanine metabolism", "phenylpropanoid biosynthesis", "cutin, suberine and wax biosynthesis", "phenylalanine, tyrosine and tryptophan biosynthesis" and "citrate cycle (TCA cycle)". Finally, we proposed an intriguing transcriptome-mediated pollen abortion and male sterility network for FA99. These findings provided data on the molecular mechanism of fertility conversion in thermo-sensitive cytoplasmic male sterility wheat.

Systematic Analysis and Identification of Drought-Responsive Genes of the CAMTA Gene Family in Wheat (Triticum aestivum L.).

The calmodulin-binding transcription activator (CAMTA) is a Ca2+/CaM-mediated transcription factor (TF) that modulates plant stress responses and development. Although the investigations of CAMTAs in various organisms revealed a broad range of functions from sensory mechanisms to physiological activities in crops, little is known about the CAMTA family in wheat (Triticum aestivum L.). Here, we systematically analyzed phylogeny, gene expansion, conserved motifs, gene structure, cis-elements, chromosomal localization, and expression patterns of CAMTA genes in wheat. We described and confirmed, via molecular evolution and functional verification analyses, two new members of the family, TaCAMTA5-B.1 and TaCAMTA5-B.2. In addition, we determined that the expression of most TaCAMTA genes responded to several abiotic stresses (drought, salt, heat, and cold) and ABA during the seedling stage, but it was mainly induced by drought stress. Our study provides considerable information about the changes in gene expression in wheat under stress, notably that drought stress-related gene expression in TaCAMTA1b-B.1 transgenic lines was significantly upregulated under drought stress. In addition to providing a comprehensive view of CAMTA genes in wheat, our results indicate that TaCAMTA1b-B.1 has a potential role in the drought stress response induced by a water deficit at the seedling stage.

Genome Wide Identification and Characterization of Wheat GH9 Genes Reveals Their Roles in Pollen Development and Anther Dehiscence.

Glycoside hydrolase family 9 (GH9) is a key member of the hydrolase family in the process of cellulose synthesis and hydrolysis, playing important roles in plant growth and development. In this study, we investigated the phenotypic characteristics and gene expression involved in pollen fertility conversion and anther dehiscence from a genomewide level. In total, 74 wheat GH9 genes (TaGH9s) were identified, which were classified into Class A, Class B and Class C and unevenly distributed on chromosomes. We also investigated the gene duplication and reveled that fragments and tandem repeats contributed to the amplification of TaGH9s. TaGH9s had abundant hormone-responsive elements and light-responsive elements, involving JA-ABA crosstalk to regulate anther development. Ten TaGH9s, which highly expressed stamen tissue, were selected to further validate their function in pollen fertility conversion and anther dehiscence. Based on the cell phenotype and the results of the scanning electron microscope at the anther dehiscence period, we found that seven TaGH9s may target miRNAs, including some known miRNAs (miR164 and miR398), regulate the level of cellulose by light and phytohormone and play important roles in pollen fertility and anther dehiscence. Finally, we proposed a hypothesis model to reveal the regulation pathway of TaGH9 on fertility conversion and anther dehiscence. Our study provides valuable insights into the GH9 family in explaining the male sterility mechanism of the wheat photo-thermo-sensitive genetic male sterile (PTGMS) line and generates useful male sterile resources for improving wheat hybrid breeding.

Integrated Analysis of Microarray, Small RNA, and Degradome Datasets Uncovers the Role of MicroRNAs in Temperature-Sensitive Genic Male Sterility in Wheat.

Temperature-sensitive genic male sterile (TGMS) line Beijing Sterility 366 (BS366) has been utilized in hybrid breeding for a long time, but the molecular mechanism underlying male sterility remains unclear. Expression arrays, small RNA, and degradome sequencing were used in this study to explore the potential role of miRNA in the cold-induced male sterility of BS366. Microspore observation showed defective cell plates in dyads and tetrads and shrunken microspores at the vacuolated stage. Differential regulation of Golgi vesicle transport, phragmoplast formation, sporopollenin biosynthesis, pollen exine formation, and lipid metabolism were observed between cold and control conditions. Pollen development was significantly represented in the 352 antagonistic miRNA-target pairs in the integrated analysis of miRNA and mRNA profiles. The specific cleavage of ARF17 and TIR1 by miR160 and miR393 were found in the cold-treated BS366 degradome, respectively. Thus, the cold-mediated miRNAs impaired cell plate formation through repression of Golgi vesicle transport and phragmoplast formation. The repressed expression of ARF17 and TIR1 impaired pollen exine formation. The results of this study will contribute to our understanding of the roles of miRNAs in male sterility in wheat.

Comprehensive analysis of formin gene family highlights candidate genes related to pollen cytoskeleton and male fertility in wheat (Triticum aestivum L.).

BACKGROUND: Formin, a highly conserved multi-domain protein, interacts with microfilaments and microtubules. Although specifically expressed formin genes in anthers are potentially significant in research on male sterility and hybrid wheat breeding, similar reports in wheat, especially in thermo-sensitive genic male sterile (TGMS) wheat, remain elusive. RESULTS: Herein, we systematically characterized the formin genes in TGMS wheat line BS366 named TaFormins (TaFHs) and predicted their functions in inducing stress response. In total, 25 TaFH genes were uncovered, majorly localized in 2A, 2B, and 2D chromosomes. According to the neighbor-joining (NJ) method, all TaFH proteins from wheat and other plants clustered in 6 sub-groups (A-F). The modeled 3D structures of TaFH1-A/B, TaFH2-A/B, TaFH3-A/B and TaFH3-B/D were validated. And different numbers of stress and hormone-responsive regulatory elements in their 1500 base pair promoter regions were contained in the TaFH genes copies. TaFHs had specific temporal and spatial expression characteristics, whereby TaFH1, TaFH4, and TaFH5 were expressed highly in the stamen of BS366. Besides, the accumulation of TaFHs was remarkably lower in a low-temperature sterile condition (Nanyang) than fertile condition (Beijing), particularly at the early stamen development stage. The pollen cytoskeleton of BS366 was abnormal in the three stages under sterile and fertile environments. Furthermore, under different stress levels, TaFHs expression could be induced by drought, salt, abscisic acid (ABA), salicylic acid (SA), methyl jasmonate (MeJA), indole-3-acetic acid (IAA), polyethylene glycol (PEG), and low temperature. Some miRNAs, including miR167, miR1120, and miR172, interacts with TaFH genes; thus, we constructed an interaction network between microRNAs, TaFHs, phytohormone responses, and distribution of cytoskeleton to reveal the regulatory association between upstream genes of TaFH family members and sterile. CONCLUSIONS: Collectively, this comprehensive analysis provides novel insights into TaFHs and miRNA resources for wheat breeding. These findings are, therefore, valuable in understanding the mechanism of TGMS fertility conversion in wheat.

Genome-wide identification and transcriptional characterization of DNA methyltransferases conferring temperature-sensitive male sterility in wheat.

BACKGROUND: DNA methyltransferase (DMT) genes contribute to plant stress responses and development by de novo establishment and subsequent maintenance of DNA methylation during replication. The photoperiod and/or temperature-sensitive genic male sterile (P/TGMS) lines play an important role in hybrid seed production of wheat. However, only a few studies have reported on the effect of DMT genes on temperature-sensitive male sterility of wheat. Although DMT genes have been investigated in some plant species, the identification and analysis of DMT genes in wheat (Triticum aestivum L.) based on genome-wide levels have not been reported. RESULTS: In this study, a detailed overview of phylogeny of 52 wheat DMT (TaDMT) genes was presented. Homoeolog retention for TaDMT genes was significantly above the average retention rate for whole-wheat genes, indicating the functional importance of many DMT homoeologs. We found that the strikingly high number of TaDMT genes resulted mainly from the significant expansion of the TaDRM subfamily. Intriguingly, all 5 paralogs belonged to the wheat DRM subfamily, and we speculated that tandem duplications might play a crucial role in the TaDRM subfamily expansion. Through the transcriptional analysis of TaDMT genes in a TGMS line BS366 and its hybrids with the other six fertile lines under sterile and fertile conditions, we concluded that TaCMT-D2, TaMET1-B1, and TaDRM-U6 might be involved in male sterility in BS366. Furthermore, a correlation analysis showed that TaMET1-B1 might negatively regulate the expression of TaRAFTIN1A, an important gene for pollen development, so we speculated regarding an epigenetic regulatory mechanism underlying the male sterility of BS366 via the interaction between TaMET1-B1 and TaRAFTIN1A. CONCLUSIONS: Our findings presented a detailed phylogenic overview of the DMT genes and could provide novel insights into the effects of DMT genes on TGMS wheat.

Comparative transcriptome and DNA methylation analysis in temperature-sensitive genic male sterile wheat BS366.

BACKGROUND: Known as the prerequisite component for the heterosis breeding system, the male sterile line determines the hybrid yield and seed purity. Therefore, a deep understanding of the mechanism and gene network that leads to male sterility is crucial. BS366, a temperature-sensitive genic male sterile (TGMS) line, is male sterile under cold conditions (12 °C with 12 h of daylight) but fertile under normal temperature (20 °C with 12 h of daylight). RESULTS: During meiosis, BS366 was defective in forming tetrads and dyads due to the abnormal cell plate. During pollen development, unusual vacuolated pollen that could not accumulate starch grains at the binucleate stage was also observed. Transcriptome analysis revealed that genes involved in the meiotic process, such as sister chromatid segregation and microtubule-based movement, were repressed, while genes involved in DNA and histone methylation were induced in BS366 under cold conditions. MethylRAD was used for reduced DNA methylation sequencing of BS366 spikes under both cold and control conditions. The differentially methylated sites (DMSs) located in the gene region were mainly involved in carbohydrate and fatty acid metabolism, lipid metabolism, and transport. Differentially expressed and methylated genes were mainly involved in cell division. CONCLUSIONS: These results indicated that the methylation of genes involved in carbon metabolism or fatty acid metabolism might contribute to male sterility in BS366 spikes, providing novel insight into the molecular mechanism of wheat male sterility.

Genomic identification and characterization of MYC family genes in wheat (Triticum aestivum L.).

BACKGROUND: MYC transcriptional factors are members of the bHLH (basic helix-loop-helix) superfamily, and play important roles in plant growth and development. Recent studies have revealed that some MYCs are involved in the crosstalk between Jasmonic acid regulatory pathway and light signaling in Arabidopsis, but such kinds of studies are rare in wheat, especially in photo-thermo-sensitive genic male sterile (PTGMS) wheat line. RESULTS: 27 non-redundant MYC gene copies, which belonged to 11 TaMYC genes, were identified in the whole genome of wheat (Chinese Spring). These gene copies were distributed on 13 different chromosomes, respectively. Based on the results of phylogenetic analysis, 27 TaMYC gene copies were clustered into group I, group III, and group IV. The identified TaMYC genes copies contained different numbers of light, stress, and hormone-responsive regulatory elements in their 1500 base pair promoter regions. Besides, we found that TaMYC3 was expressed highly in stem, TaMYC5 and TaMYC9 were expressed specially in glume, and the rest of TaMYC genes were expressed in all tissues (root, stem, leaf, pistil, stamen, and glume) of the PTGMS line BS366. Moreover, we found that TaMYC3, TaMYC7, TaMYC9, and TaMYC10 were highly sensitive to methyl jasmonate (MeJA), and other TaMYC genes responded at different levels. Furthermore, we confirmed the expression profiles of TaMYC family members under different light quality and plant hormone stimuli, and abiotic stresses. Finally, we predicted the wheat microRNAs that could interact with TaMYC family members, and built up a network to show their integrative relationships. CONCLUSIONS: This study analyzed the size and composition of the MYC gene family in wheat, and investigated stress-responsive and light quality induced expression profiles of each TaMYC gene in the PTGMS wheat line BS366. In conclusion, we obtained lots of important information of TaMYC family, and the results of this study was supposed to contribute novel insights and gene and microRNA resources for wheat breeding, especially for the improvement of PTGMS wheat lines.

Genome-wide identification and analysis of the COI gene family in wheat (Triticum aestivum L.).

BACKGROUND: COI (CORONATINE INSENSITIVE), an F-box component of the Skp1-Cullin-F-box protein (SCFCOI1) ubiquitin E3 ligase, plays important roles in the regulation of plant growth and development. Recent studies have shown that COIs are involved in pollen fertility. In this study, we identified and characterized COI genes in the wheat genome and analyzed expression patterns under abiotic stress. RESULTS: A total of 18 COI candidate sequences for 8 members of COI gene family were isolated in wheat (Triticum aestivum L.). Phylogenetic and structural analyses showed that these COI genes could be divided into seven distinct subfamilies. The COI genes showed high expression in stamens and glumes. The qRT-PCR results revealed that wheat COIs were involved in several abiotic stress responses and anther/glume dehiscence in the photoperiod-temperature sensitive genic male sterile (PTGMS) wheat line BS366. CONCLUSIONS: The structural characteristics and expression patterns of the COI gene family in wheat as well as the stress-responsive and differential tissue-specific expression profiles of each TaCOI gene were examined in PTGMS wheat line BS366. In addition, we examined SA- and MeJA-induced gene expression in the wheat anther and glume to investigate the role of COI in the JA signaling pathway, involved in the regulation of abnormal anther dehiscence in the PTGMS wheat line. The results of this study contribute novel and detailed information about the TaCOI gene family in wheat and could be used as a benchmark for future studies of the molecular mechanisms of PTGMS in other crops.

Transcriptome analysis of wheat seedling and spike tissues in the hybrid Jingmai 8 uncovered genes involved in heterosis.

MAIN CONCLUSION: Transcriptome analysis was carried out for wheat seedlings and spikes from hybrid Jingmai 8 and both inbred lines to unravel mechanisms underlying heterosis. Heterosis, known as one of the most successful strategies for increasing crop yield, has been widely exploited in plant breeding systems. Despite its great importance, the molecular mechanism underlying heterosis remains elusive. In the present study, RNA sequencing (RNA-seq) was performed on the seedling and spike tissues of the wheat (Triticum aestivum) hybrid Jingmai 8 (JM8) and its homozygous parents to unravel the underlying mechanisms of wheat heterosis. In total, 1686 and 2334 genes were identified as differentially expressed genes (DEGs) between the hybrid and the two inbred lines in seedling and spike tissues, respectively. Gene Ontology analysis revealed that DEGs from seedling tissues were significantly enriched in processes involved in photosynthesis and carbon fixation, and the majority of these DEGs expressed at a higher level in JM8 compared to both inbred lines. In addition, cell wall biogenesis and protein biosynthesis-related pathways were also significantly represented. These results confirmed that a combination of different pathways could contribute to heterosis. The DEGs between the hybrid and the two inbred progenitors from the spike tissues were significantly enriched in biological processes related to transcription, RNA biosynthesis and molecular function categories related to transcription factor activities. Furthermore, transcription factors such as NAC, ERF, and TIF-IIA were highly expressed in the hybrid JM8. These results may provide valuable insights into the molecular mechanisms underlying wheat heterosis.

Genome-wide characterization of JASMONATE-ZIM DOMAIN transcription repressors in wheat (Triticum aestivum L.).

BACKGROUND: The JASMONATE-ZIM DOMAIN (JAZ) repressor family proteins are jasmonate co-receptors and transcriptional repressor in jasmonic acid (JA) signaling pathway, and they play important roles in regulating the growth and development of plants. Recently, more and more researches on JAZ gene family are reported in many plants. Although the genome sequencing of common wheat (Triticum aestivum L.) and its relatives is complete, our knowledge about this gene family remains vacant. RESULTS: Fourteen JAZ genes were identified in the wheat genome. Structural analysis revealed that the TaJAZ proteins in wheat were as conserved as those in other plants, but had structural characteristics. By phylogenetic analysis, all JAZ proteins from wheat and other plants were clustered into 11 sub-groups (G1-G11), and TaJAZ proteins shared a high degree of similarity with some JAZ proteins from Aegliops tauschii, Brachypodium distachyon and Oryza sativa. The Ka/Ks ratios of TaJAZ genes ranged from 0.0016 to 0.6973, suggesting that the TaJAZ family had undergone purifying selection in wheat. Gene expression patterns obtained by quantitative real-time PCR (qRT-PCR) revealed differential temporal and spatial regulation of TaJAZ genes under multifarious abiotic stress treatments of high salinity, drought, cold and phytohormone. Among these, TaJAZ7, 8 and 12 were specifically expressed in the anther tissues of the thermosensitive genic male sterile (TGMS) wheat line BS366 and normal control wheat line Jing411. Compared with the gene expression patterns in the normal wheat line Jing411, TaJAZ7, 8 and 12 had different expression patterns in abnormally dehiscent anthers of BS366 at the heading stage 6, suggesting that specific up- or down-regulation of these genes might be associated with the abnormal anther dehiscence in TGMS wheat line. CONCLUSION: This study analyzed the size and composition of the JAZ gene family in wheat, and investigated stress responsive and differential tissue-specific expression profiles of each TaJAZ gene in TGMS wheat line BS366. In addition, we isolated 3 TaJAZ genes that would be more likely to be involved in the regulation of abnormal anther dehiscence in TGMS wheat line. In conclusion, the results of this study contributed some novel and detailed information about JAZ gene family in wheat, and also provided 3 potential candidate genes for improving the TGMS wheat line.

Comprehensive analyses of the annexin gene family in wheat.

BACKGROUND: Annexins are an evolutionarily conserved multigene family of calcium-dependent phospholipid binding proteins that play important roles in stress resistance and plant development. They have been relatively well characterized in model plants Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa), but nothing has been reported in hexaploid bread wheat (Triticum aestivum) and barely (Hordeum vulgare), which are the two most economically important plants. RESULTS: Based on available genomic and transcriptomic data, 25 and 11 putative annexin genes were found through in silico analysis in wheat and barley, respectively. Additionally, eight and 11 annexin genes were identified from the draft genome sequences of Triticum urartu and Aegilops tauschii, progenitor for the A and D genome of wheat, respectively. By phylogenetic analysis, annexins in these four species together with other monocots and eudicots were classified into six different orthologous groups. Pi values of each of Ann1-12 genes among T. aestivum, T. urartu, A. tauschii and H. vulgare species was very low, with the exception of Ann2 and Ann5 genes. Ann2 gene has been under positive selection, but Ann6 and Ann7 have been under purifying selection among the four species in their evolutionary histories. The nucleotide diversities of Ann1-12 genes in the four species were 0.52065, 0.59239, 0.60691 and 0.53421, respectively. No selective pressure was operated on annexin genes in the same species. Gene expression patterns obtained by real-time PCR and re-analyzing the public microarray data revealed differential temporal and spatial regulation of annexin genes in wheat under different abiotic stress conditions such as salinity, drought, cold and abscisic acid. Among those genes, TaAnn10 is specifically expressed in the anther but fails to be induced by low temperature in thermosensitive genic male sterile lines, suggesting that specific down-regulation of TaAnn10 is associated with conditional male sterility in wheat. CONCLUSIONS: This study analyzed the size and composition of the annexin gene family in wheat and barley, and investigated differential tissue-specific and stress responsive expression profiles of the gene family in wheat. These results provided significant information for understanding the diverse roles of plant annexins and opened a new avenue for functional studies of cold induced male sterility in wheat.

TaOPR2 encodes a 12-oxo-phytodienoic acid reductase involved in the biosynthesis of jasmonic acid in wheat (Triticum aestivum L.).

The 12-oxo-phytodienoic acid reductases (OPRs) are involved in the various processes of growth and development in plants, and classified into the OPRⅠ and OPRⅡ subgroups. In higher plants, only OPRⅡ subgroup genes take part in the biosynthesis of endogenous jasmonic acid. In this study, we isolated a novel OPRⅡ subgroup gene named TaOPR2 (GeneBank accession: KM216389) from the thermo-sensitive genic male sterile (TGMS) wheat cultivar BS366. TaOPR2 was predicted to encode a protein with 390 amino acids. The encoded protein contained the typical oxidored_FMN domain, the C-terminus peroxisomal-targeting signal peptide, and conserved FMN-binding sites. TaOPR2 was mapped to wheat chromosome 7B and located on peroxisome. Protein evolution analysis revealed that TaOPR2 belongs to the OPRⅡ subgroup and shares a high degree of identity with other higher plant OPR proteins. The quantitative real-time PCR results indicated that the expression of TaOPR2 is inhibited by abscisic acid (ABA), salicylic acid (SA), gibberellic acid (GA3), low temperatures and high salinity. In contrast, the expression of TaOPR2 can be induced by wounding, drought and methyl jasmonate (MeJA). Furthermore, the transcription level of TaOPR2 increased after infection with Puccinia striiformis f. sp. tritici and Puccinia recondite f. sp. tritici. TaOPR2 has NADPH-dependent oxidoreductase activity. In addition, the constitutive expression of TaOPR2 can rescue the male sterility phenotype of Arabidopsis mutant opr3. These results suggest that TaOPR2 is involved in the biosynthesis of jasmonic acid (JA) in wheat.

Genome-wide identification and evolutionary analyses of bZIP transcription factors in wheat and its relatives and expression profiles of anther development related TabZIP genes.

BACKGROUND: Among the largest and most diverse transcription factor families in plants, basic leucine zipper (bZIP) family participate in regulating various processes, including floral induction and development, stress and hormone signaling, photomorphogenesis, seed maturation and germination, and pathogen defense. Although common wheat (Triticum aestivum L.) is one of the most widely cultivated and consumed food crops in the world, there is no comprehensive analysis of bZIPs in wheat, especially those involved in anther development. Previous studies have demonstrated wheat, T. urartu, Ae. tauschii, barley and Brachypodium are evolutionarily close in Gramineae family, however, the real evolutionary relationship still remains mysterious. RESULTS: In this study, 187 bZIP family genes were comprehensively identified from current wheat genome. 98, 96 and 107 members of bZIP family were also identified from the genomes of T.urartu, Ae.tauschii and barley, respectively. Orthology analyses suggested 69.4 % of TubZIPs were orthologous to 68.8 % of AetbZIPs and wheat had many more in-paralogs in the bZIP family than its relatives. It was deduced wheat had a closer phylogenetic relationship with barley and Brachypodium than T.urartu and Ae.tauschii. bZIP proteins in wheat, T.urartu and Ae.tauschii were divided into 14 subgroups based on phylogenetic analyses. Using Affymetrix microarray data, 48 differentially expressed TabZIP genes were identified to be related to anther development from comparison between the male sterility line and the restorer line. Genes with close evolutionary relationship tended to share similar gene structures. 15 of 23 selected TabZIP genes contained LTR elements in their promoter regions. Expression of 21 among these 23 TabZIP genes were obviously responsive to low temperature. These 23 TabZIP genes all exhibited distinct tissue-specific expression pattern. Among them, 11 TabZIP genes were predominantly expressed in anther and most of them showed over-dominance expression mode in the cross combination TY806 × BS366. CONCLUSIONS: The genome-wide identification provided an overall insight of bZIP gene family in wheat and its relatives. The evolutionary relationship of wheat and its relatives was proposed based on orthology analyses. Microarray and expression analyses suggested the potential involvement of bZIP genes in anther development and facilitated selection of anther development related gene for further functional characterization.