ERK-mediated regulation of non-coding RNAs during development and disease

PROJECT SUMMARY Successful reproduction through the fusion of the sperm and oocyte is essential for the perpetuation of species. In human females, oocytes complete meiosis I at birth, and enter a long period of meiotic II arrest until onset of meiotic maturation at puberty. Because the oocytes are quiescent and arrested during this period, RNAs are loaded into the developing oocytes prior to the arrest and these RNAs are critical for early embryonic development. Mechanisms that regulate generation and protection (from degradation) of maternal RNAs during the long meiotic arrest as well as mechanisms that regulate the degradation of these RNAs in the embryo remain an active area of investigation. Our work in C. elegans and work from mammalian models in the past few years turned the light on regulation of the maternal transcriptome which dictates oocyte quality and impacts progeny development. Specifically, we uncovered a direct link between RAS/ERK growth factor signaling and the small RNA biogenesis factors Dicer1, Drosha and DIS3 (an RNA exosomal component) which regulates distinct populations of small non-coding RNAs and thus the maternal transcriptome and proteome. We propose a model wherein ERK-mediated phosphorylation of Dicer1 (and a subsequent arginine methylation of Dicer1), phosphorylation of Drosha and DIS3 results in a regulatory circuit that fine tunes the generation of small non-coding RNAs in specific subsets and regulates the maternal and zygotic transcriptome and proteome. We investigate this model in vivo during oocyte development and oocyte-to-embryo transition using a combination of live imaging, next generation sequencing, single oocyte sequencing, mass spectrometric and proteomic methods, CRISPR Cas9 genome editing and cell biological assays. We find that Dicer1, Drosha and Dis3 are phosphorylated in mammals as well. Additionally, we identified arginine methylation of Dicer1 adjacent to the phosphorylation event in mammalian cell culture system. Given their conserved role in RNA biology, reproduction and their aberrations associated with cancer onset and progression, we expect this work to have direct relevance to human biology.