The accumulation of DNA damage and mutations could lead to genomic instability and diseases such as cancer. For instance, mutations in the breast cancer associated (BRCA) 1 and 2 genes are associated with many cases of hereditary cancers including breast and/or ovarian cancer. The BRCA proteins are critical for the homologous recombination pathway, which repairs DSBs. Poly(ADP-ribose) polymerase-1 (PARP-1) is highly efficient in detecting DNA strand breaks through a DNA-break-sensing motif. Activated PARP-1 immediately modifies itself and histones with ADP-ribose polymer by binding to DNA strand breaks. The enzymatic activity of PARP-1 is suggested to facilitate DNA damage repair, and PARP-1-deficient animals and cells with PARP inhibitor show heightened sensitivity to DNA-damaging agents. Recently, inhibition of PARP-1 catalytic activity is shown to be lethal in cells lacking functional BRCA 1 or 2. However, the mechanism whereby PARP-1 enzymatic activity, including targets of the ADP-ribosylation, facilitates the DNA damage response and repair is not well characterized. Our preliminary data within this proposal show that PARP-1 modifies nucleosomal histones with ADP-ribose in response to a DNA-damaging agent, suggesting that ADP-ribosylated histones (ADPr-Hs) may play important roles in the DNA damage response. The overall hypothesis is that DNA damage-induced histone ADP-ribosylation facilitates to recruit DNA repair factors to the DNA damage sites. Major goals of this proposal are to determine the ADP-ribosylation of histones in response to DNA-damaging chemotherapeutic agents and determine DNA repair factors recruited to nucleosomes containing ADPr-Hs.
Specific Aim #1: Determine the level of ADPr-Hs in normal and repair-deficient cells in response to DNA damaging chemotherapeutic agents.
Specific Aim #2: Identify ADP-ribose-associated DNA repair factors in normal and BRCA mutant cells.