| The transmission of genetic information from parent to daughter cells is a fundamental process essential to all life. Cells have evolved multiple mechanisms to protect the fidelity of genetic information. However, perfect DNA replication and repair is not desired as it would prevent evolution and generation of diversity. Thus cells also have mechanisms to introduce mutations into the genome. Using high-throughput genomic approaches, we isolated genes involved both in DNA damage repair as well as in mutagenesis. Characterization of DOA1, a gene isolated from the DNA damage repair screen, improved our understanding of ubiquitin homeostasis under DNA damage conditions. Specifically we showed that ubiquitin trimers were broken down to monoubiquitin, in a Doa1-dependent manner, and channeled into DNA damage tolerance pathways as well as to facilitate histone H2B ubiquitination. The mutagenesis screen isolated RNR4, a component of ribonucleotide reductase, a key enzyme involved in dNTP synthesis. Surprisingly we showed that rather than facilitating lesion bypass by the conventional TLS polymerases, the upregulation of dNTP levels following DNA damage allowed a replicative polymerase, Poldelta to bypass lesions. Remarkably, we also showed that this novel mutation pathway can be efficiently inhibited with a small molecule. Moreover, the mutagenesis screen also isolated genes involved in the maintenance of ploidy. Most genes encoded components of chromatin remodeling showing just how important chromatin modification is to the faithful transmission of genetic information. Lastly, we also improved the conventional methods of measuring mutation and showed that the contribution of REV3 to mutation may be overestimated. In all, these studies have improved our understanding of how cells mutate and allowed us to begin to inhibit these processes as a therapeutic strategy. |
