DNA damage response
DNA within cells is under continual assault from metabolic and environmental sources. In response to the ensuing damage, cells activate a signaling network called the DNA damage response (DDR). Defects in this response can lead to numerous hereditary cancer syndromes and can underlie the genomic instability which is a hallmark of many sporadic cancers. The DDR promotes genomic integrity by targeting hundreds of factors in diverse pathways ranging from DNA replication and repair to cell-cycle arrest, senescence, and immune regulation. While much is known about these core pathways, the complex regulatory events coordinating them are less well understood. Our lab aims to elucidate biochemical and genetic relationships between DDR factors to understand how they are integrated and collectively regulated.
Replication stress and cancer
Replication fork collapse can induce chromosome instability and mutagenic events that cause cancer. Organisms have therefore evolved pathways to stabilize stalled replication forks and to repair collapsed forks through processes such as homologous recombination (HR). Multiple factors involved in HR and replication fork stabilization, such as BRCA1 and BRCA2, are mutated in hereditary cancer syndromes, highlighting the importance of these pathways. We have demonstrated that the ubiquitin ligase RFWD3, which is mutated in the cancer predisposition syndrome Fanconi anemia, ubiquitinates the single-stranded DNA binding protein RPA to promote homologous recombination at stalled replication forks and replication fork restart (Mol Cell 2015b). We are currently studying RFWD3 function in the replication stress response and elucidating novel mechanisms of replication fork stabilization and repair.
Quantitative proteomics in ubiquitin signaling
Execution of the DDR relies upon a dynamic array of protein modifications, with phosphorylation playing a historically central role. It is now evident that the DDR also depends on ubiquitin signaling. Numerous ubiquitin ligases have been implicated in the response, yet finding their substrates by simple binding techniques can be difficult due to weak substrate interactions. To circumvent this problem, we have pioneered a quantitative proteomic approach to globally profile ubiquitination. Initially, we used this approach to identify substrates of Cullin- RING ubiquitin ligases (Cell 2011), which are involved in numerous DNA repair processes. Subsequently, we used it to uncover novel ubiquitination events directly stimulated by DNA damage (Mol Cell 2015a), demonstrating the vast breadth of ubiquitin signaling in the DDR. We are continuing to use innovative proteomic approaches to characterize novel and poorly understood ubiquitin ligases in DNA damage signaling pathways.
Targeted cancer therapy
Defects in the DNA damage response can render tumors dependent upon specific DNA repair pathways for survival. Moreover, targeted modulation of the DDR can affect tumor sensitivity to genotoxic chemotherapy and radiation. Increased understanding of DNA repair pathways will lead to enhanced opportunities for developing therapies that target cancers with DNA repair defects, and for improving the efficacy of genotoxic treatments. We are employing methods to translate our work to the development of such therapies.