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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.

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.

The pollen fertility of photoperiod/temperature sensitive genic male sterile (P/TGMS) wheat is controlled by light and/or temperature. Circular RNA (circRNA) and long non-coding RNA (lncRNA) are known to participate in the development of anthers in plants, but their impact on male sterility in the P/TGMS line is not well understood. In this study, we carried out high-throughput sequencing to investigate the differential expression of lncRNAs and circRNAs and their biological functions in anthers of photo-thermosensitive genic male sterile (PTGMS) wheat line BS366-42L during the transition phase of male fertility under four different photoperiod and temperature treatments. Eight lncRNAs, 40 mRNAs and three circRNAs were screened out and thought as essential candidates that closely related to male sterility. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to predict the potential functions of differentially expressed RNAs. The results indicated that carbohydrate-related metabolism was important for male sterility in the wheat PTGMS line BS366-42L. lncRNA/circRNA-mRNA-miRNA (ceRNA) integrate networks were constructed to reflect their complex inner association with male sterility. Our study provides a systematic perspective on the potential function of RNAs in male fertility in PTGMS lines of wheat.

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.

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.

The miR167 family has been shown to play roles in regulating plant fertility by control the expression of a set of ARF transcription factors. In this study, the precursor of miR167a was cloned from the photo–thermo-sensitive genic male sterile (PTGMS) wheat line BS366. Moreover, we identified the target gene, TaARF8, from PTGMS wheat line BS366, and used the 5′-RACE assay to confirm the cleavage site. Overexpression (OE) of TaemiR167a in Arabidopsis induced male sterility phenotypes, and reduced the expression levels of AtARF6 and AtARF8. Meanwhile, OE of TaemiR167a reduced the content of endogenous jasmonic acid (JA) and indole acetic acid (IAA) in transgenic Arabidopsis lines. In PTGMS wheat line BS366, the expression levels of TaemiR167a were up-regulated in anther tissues during pollen development process, while the transcriptional levels of TaARF8 were down-regulated under sterile conditions. Importantly, TaemiR167a was sensitive to low temperature (10°), and was highly expressed in anther tissues of BS366 at pollen developmental stage 3 in sterile environment. Then, we measured the endogenous JA and IAA in BS366 anther tissues, and found that the accumulation of JA and IAA in anther was abnormal during the pollen development process. These results suggested that TaemiR167a-mediated regulation of TaARF8 and changes of endogenous JA and IAA during pollen developmental process took part in the pollen fertility control in PTGMS wheat line BS366.

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.

Wheat photo-thermosensitive genic male sterile (PTGMS) line is a vital material in the two-line hybrid wheat breeding system in which functional pollen production is highly associated with temperature during early developmental stage. Understanding the potential mechanism of pollen infertility induced by low temperature in PTGMS wheat is crucial for the effective utilization of genetic resources to guide wheat breeding. Herein, we combined full-length single-molecular sequencing and Illumina short reads sequencing data to obtain the high-resolution spatio-temporal transcriptome map of pollen under low temperature stress at mother cell, dyad and tetrad stages in PTGMS line BS366. Cytological descriptions and whole transcriptome analysis revealed a global landscape of low temperature altered pollen fertility transformation via regulating the transcriptional patterns of cytoskeleton-related lncRNAs and their target genes, which involved in the calcium signaling and vesicle trafficking pathways on cytoskeleton homeostasis at different stages of meiosis. Overall, our results provided the transcriptional and cytological evidences for understanding the low temperature-induced pollen sterility deficiency in PTGMS wheat line.

Male sterility is a widespread phenomenon in plants. The regulation of the pollen cytoskeleton is closely related to fertility in wheat. In this study, the distribution of the pollen cytoskeleton was observed by fluorescence staining in the conventional controlled wheat line CP806 and the genic male sterile wheat line BS366 throughout meiosis. The distribution of microfilaments and microtubules was disordered and asymmetrical in pollen cells of BS366, and the structure of the spindle body was unclear and nearly undetectable. Based on Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses, five cytoskeleton-related KEGG pathways, four GO terms, and 35 differentially expressed genes (DEGs) were identified, including eight genes assigned to GO functions and 27 genes assigned to KEGG pathways. Additionally, 86 differentially expressed transcription factors (TFs), belonging to seven TF families, were obtained. We performed qRT-PCR assays for four genes among the eight genes identified in GO enrichment analyses, eight genes among the 27 genes identified in KEGG pathway analyses, and seven TFs from seven TF families. All of these 19 candidate genes showed disparate expression patterns in CP806 and BS366, suggesting that these genes are involved in the regulation of the pollen cytoskeleton in BS366. In addition, we found lower levels of endogenous jasmonic acid (JA) and brassinolide (BR) in BS366 than that in CP806, suggesting that the biosynthetic pathways of JA and BR are abnormal in BS366. These findings provide gene resources for clarifying cytoskeleton-mediated male sterility in genic male sterile wheat lines.

Thermosensitive male sterility plays an important role in wheat fertility and production. As a key enzyme for chlorophyll degradation, pheophorbide a oxygenase (PaO) can suppress cell death in plants. We cloned the wheat gene TaPaO1 from the thermosensitive genetic male sterile (TGMS) line BS366; it encodes a typical PaO protein, containing a conserved Rieske [2Fe-2S] iron–sulphur motif, a mononuclear non-heme iron-binding motif, and a C-terminal CxxC motif. TaPaO1 was expressed in all tissues and was upregulated during the meiosis stage of BS366 anthers under low temperature. Subcellular localization of TaPaO1 specifically labelled the surrounding of chloroplasts. TaPaO1 regulated by RD29A promoter which responded to low temperature led to pollen sterility in transgenic tobacco. Expression analysis showed that TaPaO1 exhibited a higher level of expression in the anther than in other tissues in transgenic tobacco plants during low temperature treatment. We propose that the higher senescence-related activity of TaPaO1 may lead to the cell death of anthers, which happens at an early developmental stage under low temperature. These results provide new insights into the function of PaO during the early developmental stage of anthers. PaO is closely related to cell death regardless of whether it exhibits increased activity or inactive.

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.

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. Electronic supplementary material The online version of this article (10.1186/s12864-018-5116-9) contains supplementary material, which is available to authorized users.

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.

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. Electronic supplementary material The online version of this article (doi:10.1186/s12864-017-3582-0) contains supplementary material, which is available to authorized users.

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.