The Role of Double-Stranded RNA Binding Protein R2D2 and Lump in Drosophila Gametogenesis

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2010-05-14

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Sanders, Charcacia Tiana

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Abstract

In Drosophila melanogaster, fertility requires the successful development of germline cells into mature gametes. This process is dependent on multiple factors that coordinate migration, proliferation, and differentiation of germline stem cells. Previous studies have shown dsRNA binding proteins have an important role in the induction and maintenance of germ cells. In this study, I investigated the requirement of dsRNA binding proteins, R2D2 and LUMP, in Drosophila melanogaster gametogenesis. I show R2D2 functions in the ovary, specifically in the somatic tissues giving rise to the stalk and other follicle cells critical for establishing the cellular architecture of the oocyte. Most interestingly, the female fertility defects in r2d21 are dramatically enhanced when one copy of the dcr-1 gene is missing. This indicates dicer-1 and r2d21 operate in the same fertility pathway. Furthermore, Dicer-1 protein coimmunoprecipitates with R2D2 antisera, indicating these proteins function in the same protein complex. Thus, r2d21 mutants have reduced viability and defective female fertility that stems from abnormal follicle cell function and Dicer-1 impacts this process. This is the first indication R2D2 functions beyond its known role in RNA interference to include ovarian development in Drosophila. I also studied a second putative double-stranded RNA binding protein (dsRBP). I found that lump is required for male fertility and there is an absence of motile sperm in lump mutant testes squash preparations and the seminal vesicles. The early stages of spermatogenesis, including mitosis, meiosis, and cytokinesis steps are unaffected in lump mutants. This indicated lump is likely required late in sperm development. The spermatid individualization complex is disrupted consistent with an individualization-deficient phenotype. A wildtype genomic rescuing transgene was able to rescue fertility and individualization. However, a lump transgene carrying a point mutation in the first dsRNA binding domain did not rescue fertility or individualization suggesting this domain is essential for lump function. Thus, it is likely that these two dsRNA binding proteins regulate gene expression in the ovary and testes and are essential for normal fertility.

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