Cancer is the second leading cause of death in America, causing ~1 of every 4 deaths in 2009. Unrepaired DNA damage and errors during DNA replication cause mutations that contribute to the development of spontaneous cancers. In addition, most chemotherapy drugs kill cancer cells by causing DNA damage or disrupting DNA replication or repair. Replication Protein A (RPA) is a three-subunit complex that binds ssDNA non-specifically with high affinity and is essential for DNA replication, DNA repair and the cellular DNA damage responce. Null mutations in RPA are lethal and affect multiple processes. The RPA complex is composed of three subunits, RPA1, RPA2 and RPA3. Normal human tissues also contain an alternative RPA complex (aRPA) in which the RPA2 subunit is replaced by a primate-specific isoform RPA4. aRPA is present in normal tissues but down-regulated in cancer cells. Functionally, aRPA supports DNA repair, recombination and checkpoint activation but does not support chromosomal DNA replication. Our research currently focuses on understanding the functions of the two human RPA complexes, RPA and aRPA in normal cells and their role in the development of cancer and other human diseases.
My laboratory has been study cellular DNA metabolism for more than 20 years. My lab has experience with a wide-range of biochemical, molecular and cellular approaches for analyzing DNA metabolism in vitro and in vivo. We have extensive experience with purification and biochemical analysis of proteins involved in DNA replication and repair. We have either developed or applied many methods for examining DNA binding and functional analysis of RPA in DNA replication and repair. We also carried out structural analysis of RPA and have established a knockdown-reconstitution system for analyzing the function of different forms of RPA or other proteins in human cells. These studies have developed methodology for determining the functions of RPA in cellular DNA replication, DNA repair (including methods for analyzing the efficiency of specific DNA repair pathways) and checkpoint activation. My lab has generated and purified a large number of mutant forms of RPA. We have domain deletion and point mutations at disrupted the function of individual domains of RPA, a number of mutants with altered DNA binding properties and phosphorylation mimetic and un-phosphorylatible forms. We have also recently identified several separation-of-function mutations in RPA that support DNA replication but not DNA repair.