Thesis advisor: Laura Hake / The completion of meiosis in Xenopus oocytes requires the coordinated translation of stored mRNAs. CPEB, the cytoplasmic polyadenylation element binding protein, controls the translation of developmentally important early-class maternal mRNAs. Resumption of meiosis through stimulation with progesterone leads to the phosphorylation and activation of CPEB. This results in the lengthening of the poly(A) tails and translation of mRNAs containing the cytoplasmic polyadenylation element (CPE). XGef, a putative guanine nucleotide exchange factor, binds to and is required for CPEB activation. Translation of c-mos, a MAPK kinase kinase, is controlled by CPEB, and activation of the Mos/MAPK pathway is required for meiotic maturation. In addition, the synthesis of Ringo protein, an atypical cdk binding protein and activator, is required for progesterone-induced maturation, though Ringo is able to stimulate resumption of meiosis independent of progesterone. Although much work has been done to understand the key events leading to activation of maturation promoting factor (MPF) and meiotic maturation, the events immediately following progesterone stimulation remain unclear, particularly regarding the role of XGef. The work that follows describes experiments performed to further understand the role of XGef in meiotic maturation through both Ringo and MAPK activity. It was found that XGef and Ringo interact directly and form a complex throughout early meiosis. XGef is involved in Ringo’s influence during meiosis, specifically through MEK-activation of MAPK. Notably, XGef functions in a common pathway and complex with Ringo most likely to influence CPEB phosphorylation and activation. / Thesis (BS) — Boston College, 2009. / Submitted to: Boston College. College of Arts and Sciences. / Discipline: College Honors Program. / Discipline: Biology.

Thesis advisor: Laura E. Hake / Metazoan development depends on cytoplasmic polyadenylation, a key mechanism that controls the translation of maternally deposited mRNAs. In Xenopus laevis oocytes, CPEB regulates the translation of several developmentally important mRNAs, which drive meiotic progression and the production of fertilizable eggs. Most of our current knowledge of this process, also referred to as oocyte maturation, has been acquired from experiments conducted in Xenopus laevis oocytes. Despite over 30 years of research devoted to the exploration of progesterone signaling during maturation, the very early events that occur from progesterone receptor engagement to CPEB activation are not well understood. XGef, a putative Rho family guanine nucleotide exchange factor (GEF), interacts with CPEB and facilitates CPEB activation and timely meiotic progression. To further our understanding of XGef function during meiotic progression, the requirement for exchange factor activity and the activities of several Rho GTPases during maturation were examined. Despite previous reports of XGef activation of Cdc42 in mammalian cell culture, XGef does not stimulate the activation of Cdc42 in maturing Xenopus oocytes. Further, Cdc42 activity does not affect CPEB phosphorylation and overexpression of a dominant negative Cdc42 mutant does not affect maturation. Inhibition of Toxin B sensitive Rho GTPases, including Cdc42, Rac1 and Rho A-C, also fails to affect CPEB activation or meiotic progression. Lastly, the overexpression of XGef exchange deficient point mutants did not affect maturation compared to oocytes overexpressing wildtype XGef. Together, these results suggest that as a facilitator of CPEB activation and meiotic progression, XGef functions independently of exchange factor activity and Rho GTPase activation. Additionally, we found that XGef activity influences the function of RINGO/CDK1, a novel component of the progesterone signaling pathway. XGef inhibition depresses RINGO-induced GVBD, whereas XGef overexpression enhances this process. XGef interacts with RINGO in oocyte extracts and the interaction is direct in vitro. Our protein interaction data, in total, suggest that a XGef/RINGO/MAPK/CPEB complex forms in ovo to facilitate CPEB activation. Lastly, inhibition of RINGO activity directly compromises CPEB phosphorylation during early maturation, which suggests that RINGO/CDK1 directly mediates CPEB-activation. / Thesis (PhD) — Boston College, 2009. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.