The long term objectives of the proposed research are to understand the basic mechanisms of messenger RNA (mRNA) localization, and how this process is used in deployment of body patterning morphogens. Localized mRNAs have been described in animal, plant and yeast cells, where they often act in establishment or propagation of the cellular asymmetries that allow diverse cell types to be formed. Thus progress here may be of general relevance to the development of all organisms. Localization of the bicoid (bcd) mRNA to the anterior pole of the oocyte is a prerequisite for the formation of an anteroposterior gradient of Bcd protein, which patterns the anterior regions of the animal. The proposed work addresses the mechanisms underlying localization of bcd mRNA. A primary focus will be on the RNA recognition events that associate bcd mRNA with the localization machinery. The recent finding that overlapping and redundant recognition events (A and B) direct the early stages of localization provides tools - both genetic and biochemical - that will be used to better define the RNA elements involved. Related experiments will be used to search for and define RNA elements involved in secondary recognition events that direct later stages of mRNA localization. Overall, this line of investigation has two goals: facilitating the identification of binding factors; and providing insights into how complex programs of mRNA localization are directed by cis-acting regulatory elements. Proposed studies are also aimed at identifying, isolating and characterizing the factors that bind the RNA localization elements. Much of this work will focus on the factors specifically involved in the A recognition event, which is better understood than the B event. In addition, this analysis will involve further study of Ex1, a protein that acts in a secondary recognition event. Immediate goals are to identify mutants in the exl gene, which has been cloned, and to begin to characterize the contacts made by Ex1 protein. The long term goal of the work with all binding factors is to establish how RNA binding leads to RNA movement - do the RNA binding factors also bind to molecular motors, and if not, how is movement achieved? A second focus of the proposed studies is the continued development of a system to monitor mRNA movements inside the Drosophila ovary. An immediate goal is to ask if bcd mRNA sometimes moves along well-organized microtubule tracks, as is widely believed but as yet unproven.
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