Identification and Characterization of the Cdc73 Transcription Factor as a Suppressor of Genome Instability

Genome instability is an increasingly well-appreciated aspect of tumorigenesis, and genome rearrangements, such as translocations, copy number changes, and aneuploidy, are seen in many cancers. The Kolodner Laboratory has developed a variety of assays to study the formation of Gross Chromosomal Rearrangements (GCRs) in the model organism Saccharomyces cerevisiae, and our goal based on this work is to leverage the power of yeast genetics to gain insights in to the mechanisms by which increased genome instability contributes to cancer in humans. In Chapter 1, we describe collaborative efforts that used a large-scale screen to identify all the genes and pathways that interact to suppress genome instability in budding yeast. This led to the identification of 183 genes that directly suppress the accumulation of GCRs, 65 of which have not been previously identified. Bioinformatic analysis of cancer genome databases found that the human homologs of these genes are mutated in 90% and 70% of ovarian and colorectal cancers, respectively. Among the 65 novel suppressors was CDC73, a member of the Paf1 Complex that functions in transcription elongation and whose human homolog is a tumor suppressor. In Chapter 2, we investigate the mechanisms by which this gene suppresses genome instability. We demonstrate loss of CDC73 synergizes with mutations in telomere maintenance genes, and we show that increased GCRs rates are due to defects in telomerase and the accumulation of recombinogenic RNA: DNA hybrids. In Chapter 3, we expand this analysis to the rest of the Paf1 Complex members and demonstrate there is not a direct correlation between loss of complex function and an increase in GCR rate. We also defined an approximately 100 residue region of Cdc73 that is necessary and sufficient for its function and determined this region is necessary for nuclear localization and binding to Paf1. These findings on the structure and function of Cdc73 provide insights into how the human homolog functions as a tumor suppresso