The second exon of adenovirus EIA negatively modulates in vitro cell transformation, tumorigenesis and tumor metastasis. These activities of exon 2 are encoded within a short region located near the C-terminus of BIA. Mutations within this region confer a hyper-transforming phenotype on EIA. The oncogenesis restraining activities of the C-terminal region of EIA correlate with the interaction of a 48 kDa cellular nuclear protein, CtBP. CtBP is a transcriptional co-repressor. Through mutational analysis, we will determine if the oncogenesis modulating activity of CtBP is linked to its transcriptional repressor activity. CtBP also interacts with a cellular protein CtIP via a CtBP-binding motif, PLDLS. Our hypothesis predicts that interaction of CtIP with CtBP antagonizes the activities of CtBP and that the C-termninus of EIA may suppress tumorigenesis by disrupting the CtBP/CIIP complex. We hypothesize that the activity of a cellular repressor that functions through the CtBP corepressor may be altered by CtIP or E1A exon 2. We will determine the effects of overexpression of CtIP or EtA on the activity of a model human cellular repressor ZEB. CtBP is a phosphoprotein. We will identify the phosphorylation sites and investigate the role of phosphorylation on CtBP functions. In preliminary studies, we have observed that CtBP interacts with the p70 subunit of DNA-dependent protein kinase (DNA-PK). We will determine if CtBP is a target for DNA-PK. Our hypothesis predicts that an oncogenic stimulus would enhance complex formation between CtBP and CtIP. We will test this hypothesis by analyzing the CtBP/CtIP complex by coimmunoprecipitation analysis in cells expressing EtA exon 1. CtIP also interacts with pRb-family tumor suppressor proteins (pRb, p107 and p130) through an Rb-binding site (LXCXE). We hypothesize that the CR2 region ofElA promotes oncogenesis through a novel pathway (in addition to the previously known pRb/E2F pathway) by disrupting complexes of CtIP and pRb-famity proteins. We propose to investigate if the CR2 region of E1A disrupts the pRb/CtIP complex in vivo and leads to oncogenic transformation in cooperation with E2F-l. Thus, our proposed studies should illuminate how a viral oncoprotein, adenovirus EtA, modulates oncogenesis novel pathways that involve cellular proteins CtBP and CtIP.
Showing the most recent 10 out of 25 publications
