The Role of Meiotic-Stage Non-Coding RNA in the Modulation of Anther & Pollen Development in Grasses

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This project will investigate the lifecycle and functions of a class of small RNAs that support pollen development in grass flowers, specifically in anthers. The experiments focus on the biogenesis, precise localization, and quantification of these 24-nucleotide RNAs, the proteins that utilize them for anther functions, and their role in plant fertility. Prior work demonstrated that these small RNAs are required for robust male fertility under even slight temperature stress: without them anther development fails. The project will focus on maize anthers because it is easy to determine accurately what developmental stage they are in, they are easy to dissect, there are numerous developmental mutants that affect anthers for which the gene has been isolated, and because understanding male fertility is important to the production of hybrid corn seed. A more detailed understanding of the molecular basis of pollen development enables improvements in seed production, including hybrid seeds; in the grasses, hybrid corn and rice have significantly boosted world food production. Outcomes of this project could include discovering new genetic pathways for more precise control of male fertility in plants, and plants with fertility that is more resilient to environmental change. The project will utilize plant genomics and targeted mutagenesis, cutting-edge imaging techniques, bioinformatics, with a focus on developmental biology. Broader impacts of the project include training of students in plant and computational biology via continued integration with long-running and successful undergraduate and high school internship programs. The project will focus specifically on the class of 24-nt phased, secondary siRNAs, known as "24-nt phasiRNAs" that are highly enriched in meiotic-stage anthers. The work will be done in maize as the primary species for analysis because there are significant, existing genetic and molecular resources. The project will address several important questions. Do meiotic cells differ from somatic cells in the accumulation of small RNAs and their functions? Which transcription factors guide the transitions required for setting cell fate and differentiation of cell type-specific properties that include phasiRNAs? How are meiotic-stage small RNAs deployed to guide anther cell fate and differentiation? Where and when do the microRNA triggers, precursor mRNAs, and biogenesis or effector proteins accumulate? The project will characterize phenotypes and molecular genetic analysis of loss-of-function knock-out mutations that impact phasiRNAs, including mutations of the Dicer-like5 (Dcl5) gene, for which preliminary data demonstrate male sterility. Project members will be trained broadly in plant small RNA biology and genomics, cell biology and microscopy, and computational methods through their individual and group projects in the analysis of the localization, biogenesis, and biochemistry of small RNAs.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.