Development of a protein purification system for identifying human DNA repair proteins, and their functional study. (360G-Wellcome-090087_Z_09_Z)

Fanconi Anemia (FA) is a genetic disease characterized by various developmental defects and predisposition to, cancer in patients. So far, mutations in 13 genes have been identified in patients (Wang, 2007). \he corresponding proteins are recognized as FA proteins and function together in DNA inter-strand cross-link (ICL) repair, also known as the FA pathway. Several major steps in ICL repair have! been reported (Raschle, 2008), yet the critical mechanism of how ICLs are recognized in th?1 cell remains elusive, and it is unclear whether other proteins are required for the damage recognition process. We have initiated the development of a new biochemical system to identify human nuclear proteins that can bind to and recognize DNA ICL. We will further optimize this system and apply it to identify novel proteins involved in DNA ICL repair. Equipped with this tool, we are seeking to understand how the repair of ICLs is initiated in vivo. We will uncover the exact functions of the proteins, as well as their integrated role in the FA pathway and their relationship with tumorigenesis. 1) Establish a biochemical purification system and use this to identify human DNA repair proteins 2) Analysis of the FANCD2/FANCI protein complex in DNA repair 3) Functional study of FANCD2/FANCI and other identified proteins in vitro and in vivo