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The role of protein phosphorylation in biological response to genotoxie stress


Medal and 53BP1 have recently emerged as the key components of the cellular defence machinery, which guards against adverse effects of geotaxis stress. This research proposal embarks on elucidating of how phosphorylation of Medal and 53BP1 regulates genetic stability in human cells.
Task 1 aim at mapping of phosphorylation sites of Medal and 53BP1 induced by DNA damage and/or replication stress.
Task 2 includes generation of the phosphor-deficient and mimicking-mimicking mutants of Mac and 53BP1 and establishing human cell lines enabling to conditionally express these mutants after down regulation of the endogenous wild-type counterparts by RNA interference. Task 3 focuses on a comprehensive analysis of the roles of the 53BP1 and Medal phosphorylation for the kinetics and velocity of the DNA damage response using a battery of state-of-art molecular, biochemical and cell biology approaches including imaging of the key underlying biochemical reactions in live cells.
Task 4 aims at generation of specific-specific antibodies to the identified residues-residues of Medal and 53BP1. Among others, these reagents will be employed in a broad immune- photochemical survey to study the potential cancer-associated checkpoint defects. The proposed experiments are expected to significantly broaden our knowledge about the fundamental genome safeguarding mechanisms and provide tools to identify how these mechanisms are subverted in human disease.

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