Regulatory Hierarchies and Roles of Non-Coding RNAs in Maize Anthers

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PI: Blake C. Meyers (University of Delaware)CoPIs: Virginia Walbot (Stanford University) and Greg Abrams (Texas Advanced Computing Center/University of Texas - Austin) Information generated from this project will provide new insights into the regulation of developmental events in the anther, the male reproductive organ, of maize and related grasses - crops that feed much of the world. New insights and resources of broad utility should ultimately contribute to controlling plant fertility in agriculture, particularly in the cereal crops. The project builds and improves on the varied and extensive outreach and education components of the PIs and their contributions via scientific leadership to continue a strong, positive impact on local and scientific communities, and on the next generation of scientists. Implementation of the "Virtual Anther" will make the project data widely accessible while helping students and the public understand how anthers develop. An undergraduate-focused project to sequence and annotate a basal grass genome will provide invaluable data for the project and to all those employing phylogenomics analysis in flowering plants, while training the next generation of plant biologists. All project outcomes will be accessible to the research community through a project website and long-term repositories such as GEO and MaizeGDB. All biological resources such as antibodies, clones and vectors will be available upon request. Seed will be deposited and distributed long-term through the Maize Genetics Cooperative. Software for the "Virtual Anther" will be made open-source and available via the Texas Advanced Computing Center website (https://www.tacc.utexas.edu/tacc-software/).Male sterility is a common phenotype in natural species, functioning as one mechanism to promote outcrossing. Furthermore, control of pollen production by plant breeders is a fundamental technology that continues to underpin hybrid seed production in many crops. In the agricultural context, there is also a growing concern over climate disruption: both heat and cold cause male sterility in crop plants. This study seeks to more broadly understand the coordination of anther development and underlying cellular processes, including chromatin remodeling, as plant cells switch from mitosis to meiosis. The specific goals are to: (1) organize hierarchies of gene expression in individual anther cell types and (2) elucidate functions of small RNAs, particularly two unusual classes of "phased" secondary small RNAs (phasiRNAs) that are highly enriched in grass anthers. These phasiRNAs are processed from an extensive set of coordinately-expressed long non-coding RNA precursors into secondary small RNAs of 21 or 24 nucleotides. Using precisely staged maize anthers and laser microdissected cell types, the project will generate copious transcriptomic and proteomic data, organize transcriptional hierarchies of transcription factors, generate extensive data on small RNAs in a spatiotemporal context, and analyze the function and biogenesis of phasiRNAs. Genetic analysis using existing and engineered mutations will be used to test for essential roles of key transcription and phasiRNA biogenesis factors in anther development. Genetic and "omics" experiments will test new hypotheses about phasiRNA functions in anther cells and in meiosis. Most of the analysis will be conducted in maize because of its exquisite developmental regularity, the large size and ease of anther dissection, and existing molecular and genetic knowledge about maize anthers. Key insights will be evaluated in rice to distinguish general and species-specific points, with additional comparisons to other grasses. To generate new hypotheses, all data will be integrated into a "Virtual Anther" visualization system.