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.

Characterization and expression analysis of chalcone synthase gene family members suggested their roles in the male sterility of a wheat temperature-sensitive genic male sterile (TGMS) line.

Chalcone synthase (CHS), also known as the plants-specific type III polyketide synthases (PKSs), catalyzes the first key step in the biosynthesis of plant flavonoids. Flavonoids are one of the most important secondary metabolites which participate in flower pigmentation and pollen fertility. Recent reports have demonstrated the role of the CHS family in plant pollen exine formation. This study focused on the potential roles of CHS in the pollen exine formation of wheat. In the present study, a genome-wide investigation of the CHS family was carried out, and 87 CHS genes were identified in wheat. TaCHS3, TaCHS10, and TaCHS13 are wheat orthologs of Arabidopsis LESS ADHESIVE POLLEN (LAP5); TaCHS58, TaCHS64, and TaCHS67 are wheat orthologs of AtLAP6. TaCHS3, TaCHS10, and TaCHS67 showed anther-specific patterns. The expression of TaCHS3, TaCHS10, and TaCHS67 was positively co-expressed with sporopollenin biosynthetic genes, including TaCYP703A2, TaCYP704B1, TaDRL1, TaTKPR2, and TaMS2. Coincidently, the expression of TaCHS3, TaCHS10, and TaCHS67, together with those sporopollenin biosynthetic genes, were repressed at the tetrads and uninucleate stages in the temperature-sensitive genic male-sterile (TGMS) line BS366 under sterile conditions. Wheat anther-specific CHS genes might participate in the exine formation of BS366 through co-expressing with sporopollenin biosynthetic genes, which will undoubtedly provide knowledge of the roles of CHS in wheat pollen development.

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.

Genome-Wide Identification and Expression Profiling Analysis of the Long-Chain Acyl-CoA Synthetases Reveal Their Potential Roles in Wheat Male Fertility.

Long-chain acyl-CoA synthetase (LACS), responsible for the conversion of free FAs into acyl-CoAs, is involved in multiple pathways of lipid metabolism. Although LACS genes in Arabidopsis have been well characterized, no detailed information concerning this family is available for wheat. In the present study, a systematic analysis was carried out for the wheat LACS family. As a result, 30 putative TaLACSs were identified. Expression analysis revealed that 22 Takacs were expressed in wheat anthers. Two orthologs of AtLACS1, TaLACS2 and TaLACS3, were repressed at the vacuolated stage in the cold-treated BS366 (a temperature-sensitive genic male-sterile line). Thus, TaLACS2 and TaLACS3 may function like AtLACS1 in wax biosynthesis in anthers, and the repression of both genes may be correlated with the male sterility of BS366. TaLACS5 is an ortholog of AtLACS5, which was expressed exclusively in anthers. TaLACS5 was repressed in the cold-treated BS366 at the tetrad, uninucleate, and vacuolated stages. The negative correlation between TaLACS5 and TaGAMYB-B, and the MYB domain found in the promoter sequence suggested that TaLACS5 may be negatively regulated by TaGAMYB-B to participate in wheat fertility. These findings will provide a valuable foundation for the understanding of the wheat LACS gene family in male fertility.

Comparative Transcriptome Analysis Reveals Hormone Signal Transduction and Sucrose Metabolism Related Genes Involved in the Regulation of Anther Dehiscence in Photo-Thermo-Sensitive Genic Male Sterile Wheat.

Anther dehiscence is an important process to release pollen and then is a critical event in pollination. In the wheat photo-thermo-sensitive genic male sterility (PTGMS) line, pollen cannot release from anther since the anther cannot dehisce during anther dehiscence stage in a sterile condition. In this study, we carried out RNA-sequencing to analyze the transcriptome of one wheat PTGMS line BS366 during anther dehiscence under fertile and sterile conditions to explore the mechanism. We identified 6306 differentially expressed genes (DEGs). Weighted gene co-expression network analysis (WGCNA) and KEGG analysis showed that DEGs were mainly related to "hormone signal transduction pathway" and "starch and sucrose metabolism". We identified 35 and 23 DEGs related hormone signal transduction and sucrose metabolism, respectively. Compared with conventional wheat Jing411, there were some changes in the contents of hormones, including JA, IAA, BR, ABA and GA3, and sucrose, during three anther dehiscence stages in the sterile condition in BS366. We performed qRT-PCR to verify the expression levels of some critical DEGs of the hormone signaling pathway and the starch and sucrose metabolism pathway. The results showed disparate expression patterns of the critical DEGs of the hormone signaling pathway and the starch and sucrose metabolism pathway in different conditions, suggesting these genes may be involved in the regulation of the anther dehiscence in BS366. Finally, we conducted a hypothesis model to reveal the regulation pathway of hormones and sucrose on anther dehiscence. The information provided new clues to the molecular mechanisms of anther dehiscence in wheat and improved wheat hybrid breeding.

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.

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.

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 of Wheat WRKY Gene Family Reveals That TaWRKY75-A Is Referred to Drought and Salt Resistances.

It is well known that WRKY transcription factors play essential roles in plants' response to diverse stress responses, especially to drought and salt stresses. However, a full comprehensive analysis of this family in wheat is still missing. Here we used in silico analysis and identified 124 WRKY genes, including 294 homeologous copies from a high-quality reference genome of wheat (Triticum aestivum). We also found that the TaWRKY gene family did not undergo gene duplication rather than gene loss during the evolutionary process. The TaWRKY family members displayed different expression profiles under several abiotic stresses, indicating their unique functions in the mediation of particular responses. Furthermore, TaWRKY75-A was highly induced after polyethylene glycol and salt treatments. The ectopic expression of TaWRKY75-A in Arabidopsis enhanced drought and salt tolerance. A comparative transcriptome analysis demonstrated that TaWRKY75-A integrated jasmonic acid biosynthetic pathway and other potential metabolic pathways to increase drought and salt resistances in transgenic Arabidopsis. Our study provides valuable insights into the WRKY family in wheat and will generate a useful genetic resource for improving wheat breeding.

The regulatory framework of developmentally programmed cell death in floral organs: A review.

Developmentally programmed cell death (dPCD) is a tightly controlled biological process. In recent years, vital roles of dPCD on regulating floral organ growth and development have been reported. It is well known that flower is an essential organ for reproduction and a turning point of plants' life cycle. Hence, uncovering the complex molecular networks which regulates dPCD processes in floral organs is utmost important. So far, our understanding of dPCD on floral organ growth and development is just starting. Herein, we summarize the important factors that involved in the tapetal degeneration, pollen tube rupture, receptive synergid cell death, nucellar degradation, and antipodal cell degradation. Meanwhile, the known factors that involved in transmitting tract formation and self-incompatibility-induced PCD were also introduced. Furthermore, the genes that associated with anther dehiscence and petal senescence and abscission were reviewed as well. The functions of various types of factors involved in floral dPCD processes are highlighted principally. The regulatory panorama described here can provide us some insights about flower-specific dPCD process.

Identification of key genes and transcription factors in ageing Arabidopsis papilla cells by transcriptome analysis.

Programmed cell death (PCD) play essential roles in plant growth and development. Stigmatic papilla cells form an indispensable organ for plant reproduction. The lifetime of papilla cells is tightly controlled, and the developmental PCD (dPCD) process is involved in papilla cell death. Hence, papilla cell death is a good model for studying on PCD process. In this study, the dPCD signal was visualized in dying papilla cells by detecting the GUS signal of the PCD-related reporter gene BIFUNCTIONAL NUCLEASE 1 (BFN1). We found that the GUS was not expressed at young stage, but strongly expressed in papilla cells at the ageing stage, indicating the PCD process was triggered to terminate the papilla cell fate. Given this, the RNA-Seq data set, which covered the information of the whole lifespan of papilla cells, was analyzed aiming to understand which genes and pathways were involved in papilla cell death. 37 differential expressed genes (DEGs) were isolated. Moreover, the pathways related to energy production and transportation, autophagy, and plant hormone signal transduction were considered as the key pathways involved in the papilla cell death. 9 types, total of 104 transcriptional factors (TFs) were identified as well. Finally, a putative working model of papilla cell death was integrated. The findings herein will enrich the knowledge of the dPCD-mediated pathway in regulating plant organ/tissue growth, development, senescence, and death. Our study will provide some referential gene resources for studying on the dPCD in other plant organs or tissues.

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.

Uncovering Male Fertility Transition Responsive miRNA in a Wheat Photo-Thermosensitive Genic Male Sterile Line by Deep Sequencing and Degradome Analysis.

MicroRNAs (miRNAs) are endogenous small RNAs which play important negative regulatory roles at both the transcriptional and post-transcriptional levels in plants. Wheat is the most commonly cultivated plant species worldwide. In this study, RNA-seq analysis was used to examine the expression profiles of miRNA in the spikelets of photo-thermosenisitive genic male sterile (PTGMS) wheat line BS366 during male fertility transition. Through mapping on their corresponding precursors, 917-7,762 novel miRNAs were found in six libraries. Six novel miRNAs were selected for examination of their secondary structures and confirmation by stem-loop RT-PCR. In a differential expression analysis, 20, 22, and 58 known miRNAs exhibited significant differential expression between developmental stages 1 (secondary sporogenous cells had formed), 2 (all cells layers were present and mitosis had ceased), and 3 (meiotic division stage), respectively, of fertile and sterile plants. Some of these differential expressed miRNAs, such as tae-miR156, tae-miR164, tae-miR171, and tae-miR172, were shown to be associated with their targets. These targets were previously reported to be related to pollen development and/or male sterility, indicating that these miRNAs and their targets may be involved in the regulation of male fertility transition in the PTGMS wheat line BS366. Furthermore, target genes of miRNA cleavage sites were validated by degradome sequencing. In this study, a possible signal model for the miRNA-mediated signaling pathway during the process of male fertility transition in the PTGMS wheat line BS366 was developed. This study provides a new perspective for understanding the roles of miRNAs in male fertility in PTGMS lines of wheat.

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.

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.

Sequence and structure prediction of RNA-dependent RNA polymerase of lily symptomless virus isolated from L. × 'Casablanca'.

The DNA sequence of the RNA-dependent RNA polymerase (RdRp) gene of lily symptomless virus (LSV), a lily-infecting member of the genus Carlavirus, was determined from nine overlapping cDNA fragments of different sizes. The complete sequence of this RdRp gene (HM070294) consisted of 5,847 nucleotides coding for a protein of 220 kDa. It had 97-98% sequence identity with RdRps of other known isolates at both the DNA and the amino acid level. Phylogenetic analysis indicated that this RdRp (designated as RdRp-DL) was closely related to the RdRp of the Korean isolate (AM516059), as well as to the RdRps from Passiflora latent virus (PLV) and Kalanchoe latent virus (KLV) of the genus Carlavirus. Hydrophobic analysis of RdRp-DL revealed a hydrophobic N-terminus and a hydrophilic C-terminus. Helices and Loops were the major secondary structures of RdRp-DL. In addition, RdRp-DL also had three coil structures. Four conserved domains were identified: typoviral methyltransferase, RNA-dependent RNA polymerase, P-loop-containing nucleoside triphosphate hydrolases and carlavirus endopeptidase. A model of the tertiary structure predicted by I-TASSER was obtained for each of these conserved domains. This is the first report of a detailed phylogenetic analysis of LSV RdRp with those of other members of the genus Carlavirus, and the first to predict the domain structures of LSV RdRp.