Investigating the Mechanisms of Fertility Regulation by Salt-Inducible Kinase 3 in OvarianGranulosa Cells

PROJECT SUMMARY/ABSTRACT Infertility is estimated to affect 1 in 6 people worldwide, with 25% of infertility cases being attributed to ovulatory dysfunction. Despite the significant economic burden of treating ovulatory dysfunction and associated morbidities, our ability to successfully treat ovarian causes of infertility is limited by an incomplete mechanistic understanding of the development of the ovarian follicle, the functional unit of the ovary. Recently, our group was the first to describe salt-inducible kinase 3 (SIK3) as a critical determinant of fertility. Strikingly, global knockout of SIK3 in mice results in complete infertility. Our preliminary data shows that tissue-specific knockout of SIK3 in the steroidogenic granulosa cells of the ovarian follicle in a mouse model resulted in sub-fertility, a significantly reduced ovulatory response, follicle growth defects, and decreased markers of granulosa cell FSH-induced differentiation. Further, our preliminary RNA-seq data from SIK3-deficient granulosa cells in vivo showed significant alterations in lipid and glucose metabolic pathway gene expression. However, the cellular mechanisms by which SIK3 acts in the granulosa cells are currently unknown. We propose two aims to elucidate SIK3’s upstream regulation, downstream targets, and role in metabolism in FSH-induced granulosa cell differentiation. In Aim 1, I will characterize the changes in SIK3 kinase activity in response to FSH in primary mouse granulosa cells, and correlate activity to changes in SIK3 phosphorylation via phosphopeptide mapping. In Aim 2, potential upstream regulators and downstream targets of SIK3 will be identified via proximity labeling and a phospho antibody array. Finally, I will investigate whether SIK3 regulates FSH-induced differentiation via the class IIa HDACs—important SIK targets and transcriptional repressors—or other targets of interest that we identify. In Aim 3, the role of SIK3 in granulosa cell metabolism will be determined by comprehensively interrogating the effects of the loss of SIK3 on key metabolic pathways via performing metabolomics and assays measuring nutrient consumption, secretion, and utilization. Upon completion of this project, we expect to gain a foundational understanding of the mechanisms by which SIK3 regulates fertility, which may inform future work on new potential therapeutic strategies for treating infertility or developing contraceptives. This proposal was designed by my strong team of mentors and myself to broaden my expertise and toolkit, allowing me to learn, for the first time, metabolism and metabolomics, proteomics, and bioinformatics, which I hope to bring to my future career as an academic physician-scientist in pediatric endocrinology.