microRNAs (miRNAs) are an abundant class of small RNAs that derive from distinctive hairpin precursor transcripts. Their biogenesis typically involves cleavage by the Drosha and Dicer RNase III enzymes, and the resultant mature miRNAs mediate extensive networks of post-transcriptional regulation. In the course of analyzing the miRNA pathway in Drosophila, we uncovered atypical miRNAs derived from short hairpin introns that we term mirtrons. Their biogenesis substitutes a seemingly obligate processing event by the Drosha RNase III enzyme with the splicing machinery. We characterized the biogenesis and evolutionary range of mirtrons across four animal species, using biochemical, molecular, genetic, and computational methods. This work led to a series of unexpected avenues that further expanded the diversity of non- canonical miRNAs, including a hybrid pathway that marries splicing with the RNA exosome to generate hairpins that serve as Dicer substrates, as well as short hairpins that generate miRNAs in a Dicer-independent mechanism in vertebrates. In addition, this work served as a foundation of our biological studies of miRNA functions. For our renewal application, we selected several directions on understanding enzymes and mechanisms that mediate the biogenesis of mirtrons and Dicer-independent miRNAs. We present extensive unpublished data for our proposed Aims, including the discovery of hundreds of mammalian mirtrons, including a novel 3'tailed subtype in mammals, the first demonstration of Dicer-independent miRNA function in Drosophila, and establishment of in vitro assays for Dicer-independent miRNA production. These preliminary data are the basis of individual gene studies to understand detailed mechanisms, which we will extend with genomewide perspectives. In addition, we wish to relate these findings to their broader implications with respect to other small RNAs (i.e. canonical miRNAs) and discovery of novel substrates. Finally, we are concerned with relating our research to larger evolutionary questions, including the evolutionary emergence of miRNAs and of RNAi itself, and with exploiting our mechanistic knowledge to advance RNA silencing as a technological tool.
microRNAs are an extensive class of ~22 nucleotide RNAs that control the activity of messenger RNAs, the templates for protein synthesis. microRNAs dysfunction is linked to disease and cancer, while reciprocally, there is great potential for exploiting microRNAs and related regulatory RNAs as research tools and therapeutic strategies. This proposal extends our long-standing commitment to dissecting mechanisms of microRNA biogenesis by exploring new pathways that generate microRNAs and investigating enzymes that influence microRNA function.
|Wen, Jiayu; Duan, Hong; Bejarano, Fernando et al. (2015) Adaptive regulation of testis gene expression and control of male fertility by the Drosophila hairpin RNA pathway. [Corrected]. Mol Cell 57:165-78|
|Westholm, Jakub O; Miura, Pedro; Olson, Sara et al. (2014) Genome-wide analysis of drosophila circular RNAs reveals their structural and sequence properties and age-dependent neural accumulation. Cell Rep 9:1966-80|
|Yang, Jr-Shiuan; Smibert, Peter; Westholm, Jakub O et al. (2014) Intertwined pathways for Argonaute-mediated microRNA biogenesis in Drosophila. Nucleic Acids Res 42:1987-2002|
|Mohammed, Jaaved; Bortolamiol-Becet, Diane; Flynt, Alex S et al. (2014) Adaptive evolution of testis-specific, recently evolved, clustered miRNAs in Drosophila. RNA 20:1195-209|
|Brown, James B; Boley, Nathan; Eisman, Robert et al. (2014) Diversity and dynamics of the Drosophila transcriptome. Nature 512:393-9|
|Jee, David; Lai, Eric C (2014) Alteration of miRNA activity via context-specific modifications of Argonaute proteins. Trends Cell Biol 24:546-53|
|Wen, Jiayu; Mohammed, Jaaved; Bortolamiol-Becet, Diane et al. (2014) Diversity of miRNAs, siRNAs, and piRNAs across 25 Drosophila cell lines. Genome Res 24:1236-50|
|Miura, Pedro; Shenker, Sol; Andreu-Agullo, Celia et al. (2013) Widespread and extensive lengthening of 3' UTRs in the mammalian brain. Genome Res 23:812-25|
|Smibert, Peter; Yang, Jr-Shiuan; Azzam, Ghows et al. (2013) Homeostatic control of Argonaute stability by microRNA availability. Nat Struct Mol Biol 20:789-95|
|Arif, Saad; Murat, Sophie; Almudi, Isabel et al. (2013) Evolution of mir-92a underlies natural morphological variation in Drosophila melanogaster. Curr Biol 23:523-8|
Showing the most recent 10 out of 61 publications