Miiko Sokka

University of Eastern Finland, Joensuu Campus
Supervisor: Juhani Syvšoja
Funding: other
Date: 13/04/2007

Functional and structural studies of human DNA damage response protein TopBP1 and its binding partners

DNA replication and damage response mechanisms are vital for proper function of acell. This is emphasized in multicellular organisms, where failure in these mechanisms may lead to cancer predisposition. Human TopBP1 protein is an essential regulator of both mechanisms. It is a modular protein with eight conserved BRCT domains, which are believed to mediate specific binding to phosphoproteins and DNA. When DNA is damaged and DNA replication is blocked TopBP1 acts in the ATR-Chk1 cell cycle checkpoint pathway. Several lines of evidence indicate that one of its principal cell physiological outcomes is the prevention of apoptosis. Less is known about the role of TopBP1 in DNA replication. In Xenopus model organism TopBP1 participates in licensing of the replication forks.

This investigation is aimed at understanding the role of TopBP1 and its partner proteins in DNA damage response of human cultured cells. We have identified several interacting proteins of TopBP1 by yeast two-hybrid system. Our tools include immunological and molecular cloning techniques, RNA interference, chromatin immunoprecipitation (ChIP), immunoelectron microscopy, in vitro cell-free DNA replication assay techniques, and protein production and purification for structural analysis with NMR spectroscopy and X-ray crystallography.