MICRORNA EXPRESSION WITHIN PERIOVULATORY MURAL GRANULOSA CELLS
Fiedler, Stephanie Deanne
University of Kansas
Molecular & Integrative Physiology
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Normal ovarian function is an important aspect of reproductive health, and loss of this function can have drastic consequences including infertility as well as an increased risk for diseases like ovarian cancer. The ability to initiate and maintain a pregnancy is dependent on the regulation of oocyte developmental competence, ovulation and corpus luteum function as each makes an important contribution to proper ovarian function. During the periovulatory period, ovulation is initiated as a surge of LH induces granulosa cells to rapidly transform from estrogen producing follicular cells to progesterone producing luteal cells. The molecular mechanisms involved with the follicular to luteal transition have been extensively studied, and many transcriptional changes have been associated with steroidogenic and/or cell differentiation pathways. Comparatively, little attention has been given to post-transcriptional regulatory events during this period despite such events having an apparent role in maintaining the integrity of gene expression. MicroRNA (miRNA) are highly conserved, 21nt long RNA molecules that offer a way to regulate gene expression at the post-transcriptional level by binding to the 3'UTR region of a target mRNA and either preventing its translation or causing its degradation. MicroRNA have been shown to be involved in cell differentiation in other systems implying that they may also have a role in the changes that occur within granulosa cells following the LH surge. Within this thesis, I discuss several studies designed to look at miRNA expression in periovulatory granulosa cells. In addition, I describe several methods that can be used to measure levels of miRNA in their mature form as well as techniques used to begin to determine their functional relevance to the processes of ovulation and luteinization. I hypothesize that specific miRNA are induced by the LH surge, and that these miRNA are involved with regulating translation of specific target mRNA(s) within periovulatory granulosa cells. Using a mouse model, I have shown 13 miRNA to be differentially expressed at 4 hours post hCG, a critical time during the periovulatory period when many changes in gene expression are known to occur. MicroRNA-132 and miRNA-212 were found to be two of the most highly upregulated miRNA at this time point; therefore, I chose to narrow our examination to the characterization of these two miRNA and their potential mRNA targets within the periovulatory follicle. In addition to miRNA studies, I have also begun to look at the gene within which miRNA-132 and 212 reside. Experiments designed to look at the expression of this gene suggest that it is regulated by LH as well. The results I present here provide the groundwork necessary for studying the function and regulation of miRNA-132 and 212 within periovulatory follicles.
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