In an ever more industrialized environment, we are increasingly exposed to a wide range of xenobiotic compounds, many of which are DNA damaging agents that have contributed to an increasing incidence of cancer. Improving our understanding of how cells respond to and repair DNA damage will allow us to better assess genotoxic stress upon exposure to environmental agents and to develop novel molecular strategies to improve cellular responses to DNA damage. In the present proposal we focus on the mechanism responsible for breast and ovarian-specific tumor suppression by the Breast Cancer-1 susceptibility gene product (BRCA1), a key molecule involved in DNA damage signaling. We previously identified the BRCT domains of BRCA1 as phosphoserine/threonine-binding domains, and this phospho-binding function has recently been shown to be critical for BRCA1's tumor suppressor function. We will conduct a comprehensive proteome-wide screen to identify direct BRCT domain-dependent binding partners of BRCA1 and downstream targets for the ubiquitin ligase activity of BRCA1 in complex with its binding partner BARD1 in estrogen-responsive tissues. The proposed studies fit well within the long term goal of our laboratory to understand, in molecular detail, how post-translational modifications by cell signaling pathways, together with modular binding domains, regulate multiple aspects of the cellular response to DNA damage and cell cycle progression. The results of the screen should provide (1) a comprehensive unbiased identification of all BRCA1-interacting proteins in estrogen-responsive breast cells, along with determination of whether they are or are not substrates for BRCA1-BARD1 ubiquitin ligase activity;(2) the identification of potential targets and biomarkers of increased risk for DNA damage-induced carcinogenesis and (3) the creation of a novel screening approach to identify phospho-specific protein-protein interactions and E3 ubiquitin ligase substrates in a genome-wide manner. Since downstream targets are likely to be misregulated in cells lacking normal BRCA1 function, our screen is expected to reveal novel targets for drug development and novel biomarkers for assessing cellular responses to genotoxic stress.
BRCA1 is a tumor suppressor protein that plays a critically important role in the DNA damage response, but the mechanism by which BRCA1 prevents tumor formation is largely unknown. Mutations in BRCA1 causes dramatically increased risk for cancer in estrogen-responsive tissues, particularly the breast and ovary. In this proposal we develop a novel proteome-wide method for comprehensively identifying the binding partners and substrates of BRCA1 in order to better understand its anti-cancer function at the molecular level.
| Morton, Stephen W; Lee, Michael J; Deng, Zhou J et al. (2014) A nanoparticle-based combination chemotherapy delivery system for enhanced tumor killing by dynamic rewiring of signaling pathways. Sci Signal 7:ra44 |
| Reinhardt, H Christian; Yaffe, Michael B (2013) Phospho-Ser/Thr-binding domains: navigating the cell cycle and DNA damage response. Nat Rev Mol Cell Biol 14:563-80 |
