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 plants, CCCH zinc finger proteins involved in secondary wall formation and anther development are poorly understood. We have functionally identified two homologous genes C3H14 and C3H15 and found that the two genes differentially regulate secondary wall formation and anther development. C3H14 contributes more to secondary wall thickening, whereas, C3H15 is more important for anther development. We performed microarray analyses on C3H14/15 overexpressors and c3h14c3h15 double mutant to to ientify differentially-expressed genes involved in the two developmental processes.
Project description:Single-cell genomics provides unprecedented potential for research on plant development and environmental responses. Here, we introduce a generic procedure for plant nuclei isolation combined with nanowell-based library preparation. Our method enables the transcriptome analysis of thousands of individual plant nuclei. It serves as alternative to the use of protoplast isolation, which is currently the standard methodology for plant single-cell genomics, although it can be challenging for some plant tissues. We show the applicability of our nuclei isolation method by using different plant materials from different species. The potential of our snRNA-seq method is shown through the characterization of transcriptomes of seedlings and developing flowers from Arabidopsis thaliana. We evaluated the transcriptome dynamics during the early stages of anther development, identified stage-specific activities of transcription factors underlying this process and predicted potential target genes of these transcription factors. Our nuclei isolation procedure can be applied in different plant species and tissues, thus expanding the toolkit for plant single-cell genomics experiments.
