The long term goal of this proposal is to aide in elucidating nucleolin's role in the DNA damage response. Our laboratory has found nucleolin relocalization from the nucleolus to the nucleoplasm following various celluar stresses. This event has been correlated with a change in the phosphorylation status of nucleolin. The study is aimed at exploring how nucleolin phosphorylation is regulated in response to genotoxic stress. There are four specific aims of the nucleolin phosphorylation studies. First, identify the sites on nucleolin whose phosphorylation changes in response to various genotoxic stresses such as ionizing radiation, camptothecin, UV and heat shock. Second, identify the protein kinase(s) and other interacting proteins responsible for the phospho-modifications by in vitro phosphorylational and mutational analysis. Third, characterize the mechanism regulating the functional interaction of the CK2 with nucleolin using immunoprecipitation techniques. And fourth, understand the functional consequences of the nucleolin phosphorylation event by studying how the phosphorylation status of nucleolin affects its interaction with DNA damage response agents such as p53, and hRPA. ? ? ? ?
