Small noncoding RNAs are central components ofthe gene regulatory circuit in organisms. For proper health and development, organisms employ protein complexes containing small endogenous RNAs to control thousands of other genes through silencing mechanism. Genetics in model organisms implicate a link between small RNAs, Argonaute proteins and processes that regulate chromatin structure, suggesting that RNA interference ^RNAi) pathways can impact large domains of the genome. Our search for an analogous link in mammals directed us to purify from rat testes the piRNA Complex (piRC), which consists at least of mammalian Piwi and the Piwi-interacting RNAs. Fiwi-interacting RNAs (piRNAs) are a diverse class of small RNAs that are longer than microRNAs and derive from intergenic clusters of between 1-lOOkb long. Piwi, a subclade of the Argonaute family, is essential for fertility in flies and male mice, and has been genetically implicated in silencing of transposable elements. Despite these features, a rigorous understanding of piRC's molecular function is still lacking. This proposal seeks to dissect the molecular function ofthe piRNAs and piRC through these specific aims: 1) Functional analysis ofthe piRNA Complex;2) Regulation of piRNA cluster transcription;3) Understanding the Genesis of piRNAs. We will apply our current biochemical techniques to probe piRC's functional activity, and develop genetic and cell biology methodologies to understand the role of piRNA clusters. Piwi proteins have been shown to be essential for proper germ cell development in animals, including mammals. However, the mechanistic understanding and gene regulatory potential ofthis ribonucleoprotein is still uncharted. This research project aims to develop systems that will drive progress in a subject relevant to child health and human development.
This application seeks funds that will facilitate investigations ofthis new mechanism for RNAi in animals and describes a comprehensive plan that will develop the career of the candidate towards building an independent research program. A deeper understanding of piRC's function will yield insight to fundamental gene regulatory processes that impact human development and fertility.
|Clark, Josef P; Rahman, Reazur; Yang, Nachen et al. (2017) Drosophila PAF1 Modulates PIWI/piRNA Silencing Capacity. Curr Biol 27:2718-2726.e4|
|Toombs, James A; Sytnikova, Yuliya A; Chirn, Gung-Wei et al. (2017) Xenopus Piwi proteins interact with a broad proportion of the oocyte transcriptome. RNA 23:504-520|
|Chirn, Gung-Wei; Rahman, Reazur; Sytnikova, Yuliya A et al. (2015) Conserved piRNA Expression from a Distinct Set of piRNA Cluster Loci in Eutherian Mammals. PLoS Genet 11:e1005652|
|Rahman, Reazur; Chirn, Gung-wei; Kanodia, Abhay et al. (2015) Unique transposon landscapes are pervasive across Drosophila melanogaster genomes. Nucleic Acids Res 43:10655-72|
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|Sytnikova, Yuliya A; Rahman, Reazur; Chirn, Gung-Wei et al. (2014) Transposable element dynamics and PIWI regulation impacts lncRNA and gene expression diversity in Drosophila ovarian cell cultures. Genome Res 24:1977-90|
|Ow, Maria C; Lau, Nelson C; Hall, Sarah E (2014) Small RNA library cloning procedure for deep sequencing of specific endogenous siRNA classes in Caenorhabditis elegans. Methods Mol Biol 1173:59-70|
|Ghiretti, Amy E; Moore, Anna R; Brenner, Rebecca G et al. (2014) Rem2 is an activity-dependent negative regulator of dendritic complexity in vivo. J Neurosci 34:392-407|
|Arnold, Cosmas D; Gerlach, Daniel; Spies, Daniel et al. (2014) Quantitative genome-wide enhancer activity maps for five Drosophila species show functional enhancer conservation and turnover during cis-regulatory evolution. Nat Genet 46:685-92|
|Clark, Josef P; Lau, Nelson C (2014) Piwi Proteins and piRNAs step onto the systems biology stage. Adv Exp Med Biol 825:159-97|
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