Functional analysis of gene expression regulation by Rb/E2F in the Caenorhabditis elegans germ line

The role of the retinoblastoma protein (RB) and its association to the sequence specific transcription factor E2F in mammalian cell culture studies have been clearly demonstrated in the regulation of the cell cycle and cell proliferation. A physical interaction between RB and E2F results in transcriptional repression, while free E2F functions as a transcriptional activator to promote cell proliferation. However, the increased tumorigenesis and differentiation defects observed in individual E2F knock out mice have suggested that the in vivo role of E2F is more complex and is not limited to cell proliferation. The research presented in this thesis demonstrates that C. elegans EFL-1(E2F) and DPL-1(DP) act together independently of LIN-35(Rb) in the germ line to promote differentiation of oocytes and embryos, a model that is compatible with the in vivo mouse phenotype. Specifically, while LIN-35(Rb) appears to act as a repressor of a diverse range of genes, EFL-1 and DPL-1 promote activation of genes that are required during oocyte differentiation and embryo development. Thus, this work presents a novel in vivo role for E2F; E2F promotes pro-differentiation genes required for fertility. I have further characterized the sterility phenotype of dpl-1 null mutants and found that its ovulation is defective due to abnormal spermatheca dilation. Providing exogenous DPL-1 in the germ line results in near complete rescue, suggesting that DPL-1 acts in the germ line to regulate spermatheca dilation. A RNAi screen of dpl-1 downstream target genes and subsequent transgene rescue experiments revealed that ovulation control by DPL-1 is primarily mediated by activity of RME-2. Thus, the results of this study indicate that C. elegans E2F acts to promote differentiation through direct activation of target genes and its role in the cell cycle may be secondary or absent in these tissues in vivo.