RNA Localization in Xenopus Ooctytes

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DESCRIPTION (provided by applicant): Establishment of polarity in the egg can be viewed as the earliest step in embryonic patterning. Thus, differences in cell fate among the early cleavage cells are a consequence of asymmetric distributions of informational molecules in the egg cytoplasm before fertilization, the basis for such polarity can be provided by localized maternal determinants in the form of mRNA. While there is evidence for the existence of localized determinants in many systems, the localization process is only now being unraveled. Among vertebrates, Vg1 mRNA is a prominent example of a localized mRNA that is thought to play a role in patterning. Vg1 mRNA encodes a growth factor-like molecule, and is localized during oogenesis to the vegetal cytoplasm of Xenopus oocytes. Restricted expression of Vgl protein in the vegetal hemisphere of the egg appears to be critical for correct patterning of the embryo, making Iocalization of Vg1 mRNA an important model for understanding how maternal molecules are localized to influence pattern and polarity. The goal of this research project is to investigate how RNA molecules can be targeted to specific regions of the cell cytoplasm to generate spatially restricted gene expression. The foundation for this investigation has been laid by our recent work studying the interactions between trans-acting localization factors and essential targeting sequences within Vg1 mRNA. We have identified and characterized key components of the localization machinery and defined consensus sequence elements within an RNA targeting signal. Moreover, we have discovered that the localization pathway initiates in the nucleus and we have uncovered distinct nuclear and cytoplasmic steps in the localization pathway. We now seek to extend these findings to elucidate the molecular mechanisms responsible for localized expression of Vg1 RNA with the following specific aims: I) To analyze nuclear events leading to cytoplasmic RNA localization. II) To characterize assembly and transport of the cytoplasmic RNP complex. III) To probe coordination of mRNA localization with other post-transcriptional control points. The proposed research is designed to provide mechanistic insight into how developmental signals are spatially distributed in the vertebrate embryo.