Project description:Our results show that ICE1 controls plant male fertility via impacting anther dehydration. The loss-of-function mutation in ICE1 gene in Arabidopsis caused anther indehiscence and decreased pollen viability as well as germination rate. Further analysis revealed that the anthers in the mutant of ICE1 (ice1-2) had the structure of stomium, though the epidermis did not shrink to dehisce. The anther indehiscence and influenced pollen viability as well as germination in ice1-2 were due to abnormal anther dehydration, for most of anthers dehisced with drought treatment and pollen grains from those dehydrated anthers had similar viability and germination rates compared with wild type. Accordingly, the sterility of ice1-2 could be rescued by ambient dehydration treatments. Likewise, the stomatal differentiation of ice1-2 anther epidermis was disrupted in a different manner compared with that in leaves. ICE1 specifically bound to MYC-recognition elements in the promoter of FAMA, a key regulator of guard cell differentiation, to activate FAMA expression. Transcriptome profiling in the anther tissues further exhibited ICE1-modulated genes associated with water transport and ion exchange in the anther. Together, this work reveals the key role of ICE1 in male fertility control and establishes a regulatory network mediated by ICE1 for stomata development and water movement in the anther.
Project description:In flowering plants, the male gametophyte, the pollen, develops in the anther. Complex patterns of gene expression in both the gametophytic and sporophytic tissues of the anther regulate this process. The gene expression profiles of the microspore/pollen and the sporophytic tapetum are of particular interest. In this study, a microarray technique combined with laser microdissection (44K LM-microarray) was developed and used to characterize separately the transcriptomes of the microspore/pollen and tapetum in rice. Expression profiles of 11 known tapetum specific-genes were consistent with previous reports. Based on the spatiotemporal expression patterns and gene ontology (GO) categories of anther-expressed genes, some noteworthy expression patterns are discussed in connection with various important biological events of anther development.
Project description:Background: Pepper (Capsicum annuum L.) is a major cash crop throughout the world. Male sterility is an important characteristic in crop species that leads to a failure to produce functional pollen, and it has crucial roles in agricultural breeding and the utilization of heterosis. Objectives: In this study, we identified many crucial factors and important components in metabolic pathways in anther and pollen development, and elucidated the molecular mechanism related to pollen abortion in pepper. Methods: Pepper pollen was observed at different stages to detect the characteristics associated with male sterility and fertility. The phytohormone and oxidoreductase activities were detected in spectrophotometric and redox reaction assays, respectively. Proteins were extracted from male sterile and fertile pepper lines, and identified by TMT/iTRAQ (Tandem mass tags/isobaric tags for relative and absolute quantitation) and LC-MS/MS (liquid chromatograph-mass spectrometer) analysis. Differentially abundant proteins (DAPs) were analyzed based on Gene Ontology annotations and the Kyoto Encyclopedia of Genes and Genomes database according to |fold change)| > 1.3 and P value < 0.05. DAPs were quantified in the meiosis, tetrad, and binucleate stages by parallel reaction monitoring (PRM). Results: In this study, we screened and identified one male sterile pepper line with abnormal cytological characteristics in terms of pollen development. The peroxidase and catalase enzyme activities were significantly reduced and increased, respectively, in the male sterile line compared with the male fertile line. Phytohormone analysis demonstrated that the gibberellin, jasmonic acid, and auxin contents changed by different extents in the male sterile pepper line. Proteome analysis screened 1645 DAPs in six clusters, which were mainly associated with the chloroplast and cytoplasm based on their similar expression levels. According to proteome analysis, 45 DAPs were quantitatively identified in the meiosis, tetrad, and binucleate stages by PRM, which were related to monoterpenoid biosynthesis, and starch and sucrose metabolism pathways. Conclusions: We screened 1645 DAPs by proteomic analysis and 45 DAPs were related to anther and pollen development in a male sterile pepper line. In addition, the activities of peroxidase and catalase as well as the abundances of phytohormones such as gibberellin, jasmonic acid, and auxin were related to male sterility. The results obtained in this study provide insights into the molecular mechanism responsible for male sterility and fertility in pepper.
Project description:Formation of functional pollen and successful fertilisation relies upon the spatial and temporal regulation of anther and pollen development. This process responds to environmental cues to maintain optimal fertility despite climatic changes. Arabidopsis transcription factors bHLH10,-89,-91 were previously thought to be functionally redundant in their control of male reproductive development, however here we show that they play distinct roles in the integration of light signals to maintain pollen development under different environmental conditions. Combinations of the double and triple bHLH10,-89,-91 mutants were analysed under normal (200μmol/m2/s) and low (50μmol/m2/s) light conditions to determine the impact on fertility. Transcriptomic analysis of a new conditionally sterile bhlh89,91 double mutant shows differential regulation of genes related to sexual reproduction, hormone signal transduction and lipid storage and metabolism under low-light. Here we have shown that bHLH89 and bHLH91 play a role in regulating fertility in response to light, suggesting they function in mitigating environmental variation to ensure fertility is maintained under environmental stress.
Project description:The CLAVATA3/ESR-RELATED (CLE) peptide hormones are required for numerous plant growth and developmental processes. However, little is known regarding the function and working mechanism of the CLEs in the anther. Here, using RNA in situ hybridization analyses, we identified 7 CLE genes that are specifically expressed in the tapetum and microsporocytes in the anther, and the dominant-negative mutant plants of each of these genes exhibited significantly reduced anther size, pollen number, and abnormal pollen wall formation. Further transcriptomic and proteomic studies on cle19, DN-CLE19, and CLE19-OX mutant lines revealed that CLE19 affected the expression of more than 1,000 genes at the RNA level and 595 at the protein level, including genes involved in pollen coat and pollen exine formation, lipid metabolism, pollen germination, and hormone metabolism processes. Phenotypic analyses of mutants of the CLE19 downstream genes GRP20, ACOS5 and MEE48 revealed that the formation of pollen exine was affected in these mutants, confirming that these genes function downstream of CLE19 in the regulation of pollen wall formation. These findings demonstrate the function and downstream genes of CLE19 and redundant genes, providing insights into working pathways of the peptide hormones in pollen development.
Project description:In flowering plants, the male gametophyte, the pollen, develops in the anther. Complex patterns of gene expression in both the gametophytic and sporophytic tissues of the anther regulate this process. The gene expression profiles of the microspore/pollen and the sporophytic tapetum are of particular interest. In this study, a microarray technique combined with laser microdissection (44K LM-microarray) was developed and used to characterize separately the transcriptomes of the microspore/pollen and tapetum in rice. Expression profiles of 11 known tapetum specific-genes were consistent with previous reports. Based on the spatiotemporal expression patterns and gene ontology (GO) categories of anther-expressed genes, some noteworthy expression patterns are discussed in connection with various important biological events of anther development. The separated transcriptomes of rice microspore/pollen and tapetum were measured at the premeiosis, meiosis, tetrad, uninuclear, bicellular, and tricelluar stages by using laser microdissection (LM)-mediated microarray.
Project description:Transcriptomic analysis of single, double and triple mutant anthers of bhlh010, bhlh089 and bhlh091. We examine here three recently duplicated Arabidopsis bHLH genes, bHLH010, bHLH089 and bHLH091, using evolutionary, genetic, morphological and transcriptomic approaches, and uncover their redundant functions in anther development. These three genes are relatively highly expressed in the tapetum of the Arabidopsis anther; single mutants at each of the bHLH010, bHLH089 and bHLH091 loci are developmentally normal, but the various double and triple combinations progressively exhibit increasingly defective anther phenotypes (abnormal tapetum morphology, delayed callose degeneration, and aborted pollen development), indicating their redundant functions in male fertility. Note: Samples in SRA were assigned the same sample accession. This is incorrect as there are different samples, hence “Source Name” was replaced with new values. Comment[ENA_SAMPLE] contains the original SRA sample accessions.
Project description:Illumina sequencing was employed to examine the expression profiles of rice anther miRNAs from the a non-pollen male sterile line Wuxiang S (WXS), one of photo-thermo sensitive genical male sterile (PTGMS) line rice, during in the fertility transition stage. A total of 493 known miRNAs and 273 novel miRNAs were identified during rice anther development. Based on the number of sequencing reads, a total of 26 miRNAs were discovered to be significant difference expression between WXS(S, Sterility) and WXS(F, Fertility), and the results were partially validated by qRT-PCR. Among these, 11 miRNAs were decreased and 15 miRNAs were increased in WXS(S) compared with WXS(F). The expression patterns for targets of osa-miR156a-j, osa-miR3879, osa-miR159c/d/e, osa-miR171a/c/e/i, osa-miR398b, osa-miR164d, osa-miR528 and osa-miR408 were selectively examined, and the results showed that there was a negative correlation on the expression patterns between miRNAs and their targets. These targets have previously been reported to be related with pollen development and male sterility, suggesting that miRNAs might act as regulators of rice anthers. Furthermore, miRNA editing events were observed. The U-to-C and U-to-A editing phenomenon was validated by molecular cloning and sequencing. Examine small RNA profiles change of four tissues of the rice non-pollen male sterile line Wuxiang S under two different environments.
Project description:Anther development is a complex process, and the study of its molecular mechanism has an important impact on plant breeding. This study aims to identify microRNA (miRNA), mRNA, long non-coding RNA (lncRNA), and circular RNA (circRNA) related to anther development of Chinese cabbage, so as to construct competitive endogenous RNA (ceRNA) regulatory networks and provide valuable knowledge for the exploration of pollen development mechanism of Chinese cabbage. A total of 9055 mRNA, 585 miRNA, 1344 lncRNA, and 165 circRNA were identified as differentially expressed in the anther of Chinese cabbage compared with Mix (roots, stems and leaves) by whole-transcriptome sequencing. The anther-related ceRNA-miRNA-target gene regulatory network through miRNA targeting relationships was constructed and 450 pairs of ceRNA relationships, including 97 DEmiRNA-DEmRNA, 281 DEmiRNA-DElncRNA, and 23 DEmiRNA-DEcircRNA interactions were obtained in Chinese cabbage. The genes in the ceRNA network were enriched in the pathways including starch and sucrose metabolism, carbon metabolism, pyruvate metabolism and carbon fixation in photosynthetic organisms, plant hormone signal transduction, and RNA degradation. This study identified some important genes and their interaction lncRNAs, circRNAs, and miRNAs involved in microsporogenesis (BraA06g035480.3C), tapetum and callose layer development (BraA09g009280.3C, BraA04g028920.3C, and BraA10g022680.3C etc), pollen wall formation (BraA06g000980.3C, BraA02g023130.3C, and BraA10g029650.3C etc), and anther dehiscence (BraA10g027200.3C, BraA04g023740.3C, and BraA04g030860.3C etc). Additionally, we analyzed the promoter activity of six anther predominant expression genes, and the results showed that they were all expressed specifically in the anther of Chinese cabbage. This study lay the foundation for further research on the molecular mechanism of anther growth and development.