Prokaryotes and eukaryotes are endowed with multiple specialized DMApolymerases that support DMA synthesis across sites of DNA damage that arrest normal high-fidelity replication. This process is called translesion DNA synthesis (TLS). This function is fundamental to the survival of cells that have suffered arrested DNA replication due to the presence of unrepaired DNA damage, and to the genesis of mutations in living cells. These polymerases may also play a role in somatic hypermutation (SH) in the immune system by filling in gaps in DNA in an error-prone manner, increasing the diversity of antibodies. Understanding the molecular mechanism of TLS is therefore central to understanding all human diseases associated with mutagenesis, especially cancer, and to understanding SH in the immune system. We have identified a polymerase, POLK, and created a mouse model for this damage inducible polymerase. The progeny of Polk- /- mice manifest a mutator phenotype and cells from these mice are sensitive to killing by UV radiation and to the polycyclic aromatic mutagen and carcinogen benzo[a]pyrene.
Specific aim 1 of this renewal proposal is to characterize the molecular basis of the mutator phenotype identified in progeny of Polk-/- mice by using mutation detection systems to clarify the in vivo functions of POLK. These systems will identify changes found in the DNA of mice lacking this enzyme.
Aim 2 is to identify the functions of the multiple POLK isoforms present in mouse testis, their biochemical interactions with other proteins and DNA as well as their ability to add nucleotide opposite DNA lesions. We have also identified proteins that interact with POLK and are pursuing how these interactions affect damage recognition and repair.
Aim 3 is to elucidate the molecular mechanism(s) of DNA polymerase recruitment during TLS through the detailed characterization of REV1 protein in mammalian cells and in the yeast S. cerevisiae, a genetically tractable model eukaryote. The results of these studies will clarify the in vivo functions of POLK and lead to further understanding of how POLK and it's protein isoforms interact with REV1L during polymerase switching will help define how this aspect of genome maintenance works to prevent mutations that lead to cancer and other diseases.
| Singer, William D; Osimiri, Lindsey C; Friedberg, Errol C (2013) Increased dietary cholesterol promotes enhanced mutagenesis in DNA polymerase kappa-deficient mice. DNA Repair (Amst) 12:817-23 |
| Guo, Caixia; Tang, Tie-Shan; Friedberg, Errol C (2010) SnapShot: nucleotide excision repair. Cell 140:754-4.e1 |
| Stancel, J Nicole Kosarek; McDaniel, Lisa D; Velasco, Susana et al. (2009) Polk mutant mice have a spontaneous mutator phenotype. DNA Repair (Amst) 8:1355-62 |
| Moore, Destaye M; Karlin, Justin; González-Barrera, Sergio et al. (2009) Rad10 exhibits lesion-dependent genetic requirements for recruitment to DNA double-strand breaks in Saccharomyces cerevisiae. Nucleic Acids Res 37:6429-38 |
| Guo, Caixia; Kosarek-Stancel, J Nicole; Tang, Tie-Shan et al. (2009) Y-family DNA polymerases in mammalian cells. Cell Mol Life Sci 66:2363-81 |
| Guo, Caixia; Tang, Tie-Shan; Bienko, Marzena et al. (2008) Requirements for the interaction of mouse Polkappa with ubiquitin and its biological significance. J Biol Chem 283:4658-64 |
| Kosarek, J Nicole; Woodruff, Rachel V; Rivera-Begeman, Amanda et al. (2008) Comparative analysis of in vivo interactions between Rev1 protein and other Y-family DNA polymerases in animals and yeasts. DNA Repair (Amst) 7:439-51 |
| Guo, Caixia; Sonoda, Eiichiro; Tang, Tie-Shan et al. (2006) REV1 protein interacts with PCNA: significance of the REV1 BRCT domain in vitro and in vivo. Mol Cell 23:265-71 |
| Guo, Caixia; Tang, Tie-Shan; Bienko, Marzena et al. (2006) Ubiquitin-binding motifs in REV1 protein are required for its role in the tolerance of DNA damage. Mol Cell Biol 26:8892-900 |
| Guo, Caixia; Fischhaber, Paula L; Luk-Paszyc, Margaret J et al. (2003) Mouse Rev1 protein interacts with multiple DNA polymerases involved in translesion DNA synthesis. EMBO J 22:6621-30 |
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