Functional characterization of the role of Imp, a Drosophila mRNA binding protein, during oogenesis
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Establishment of cell polarity requires the involvement of several posttranscriptional regulatory mechanisms, including mRNA localization and translational control. A family of highly conserved RNA binding proteins in vertebrates, VICKZ (V̲g1RBP/V̲era, I̲MP-1, 2, 3, C̲RD-BP, K̲OC, Z̲BP-1) proteins, has been shown to act in these two processes. Previous studies of the posttranscriptional mechanisms mediated by VICKZ family members have been largely limited by the lack of genetic approaches in certain vertebrate systems. Identification of Imp, the Drosophila member of the VICKZ family, opened the possibility to use genetic approaches to investigate the roles of a VICKZ family member in mRNA localization and translational control. In this dissertation, we show that Imp is associated with Squid and Hrp48, two heterogeneous proteins (hnRNP) that complex with one another to regulate localized expression of gurken (grk). In addition, Imp binds grk mRNA with high affinity in vitro and is concentrated at the site of grk localization in midstage oocytes. Mutation of the Imp gene does not substantially alter grk expression, but does partially suppress the grk mis-expression phenotype of fs(1)k10 mutants. In contrast, overexpression of Imp in germ line cells results in mislocalization of grk mRNA and protein. The opposing effects of reduced and elevated Imp activities on grk expression suggest that Imp acts in regulation of grk expression, but in a redundant way. To further explore the mechanisms by which localized expression of grk is regulated by Imp, a deficiency screen was conducted to search for dominant modifiers of the dorsalized phenotype resulting from Imp overexpression. Twelve genomic regions were identified to contain dominant modifiers of the Imp overexpression phenotype. Further characterization of mutants of genes within these genomic regions led to identification of five modifiers, including cyclin E (cycE), E2f transcriptional factor 1 (E2f1), lingerer (lig), snail (sna) and mushroom body expressed (mub). E2f1 encodes a transcriptional factor that is involved in regulating the G1 to S phase transition during mitosis. Mutation of E2f1 results in altered grk mRNA and protein distribution within oocyte, revealing a role for this gene in regulation of grk expression.