Current Research

Understanding DNA Mismatch Repair
DNA mismatch repair (MMR) detects and acts on mispaired bases.  When acting on mispaired bases in newly replicated DNA, MMR prevents the accumulation of mutations.  When acting on mispaired bases in DNA recombination intermediates, MMR prevents recombination between divergent, repeated DNA sequences and can also repair mispairs present in recombination intermediates.  Our laboratory: (1) discovered or co-discovered most of the eukaryotic MMR genes, (2) demonstrated that inherited defects in key MMR genes cause a hereditary cancer predisposition syndrome (Lynch syndrome, MIM #120435), and (3) elucidated the major cause of MMR defects in sporadic cancers.  This work provided the first demonstration that defects in a DNA repair pathway that repairs DNA damage due to endogenous metabolic errors can lead to the development of common cancers.  Our current efforts study the mechanisms of MMR reactions reconstituted in vitro with purified S. cerevisiae proteins; these studies are guided by and often prompt additional genetic analyses of S. cerevisiae MMR.  Major questions about MMR that we are studying include: 1) how is MMR coupled to DNA replication, 2) how is MMR targeted to the newly synthesized DNA strand, 3) how do exonuclease 1-dependent and exonuclease-independent MMR function, and 4) how do protein-protein and protein-mispaired base interactions promote MMR reactions.
Understanding Pathways that Prevent Genome Rearrangements
Our laboratory has worked extensively on identifying the pathways that prevent genome rearrangements.  Using S. cerevisiae as a model system, we developed the first quantitative genetic assays for measuring the rates of accumulation of gross chromosomal rearrangements (GCRs), including translocations, deletions, inversions and broken chromosomes healed by de novo telomere addition.  This critical advance allowed us to identify the first genes known to suppress GCRs and, through study of these first genes, obtain many insights into how GCRs are prevented and the mechanisms that form GCRs.  We recently extended these studies by performing large-scale genetic screens in S. cerevisiae using multiple GCR assays to identify all of the GCR-suppressing (GIS) genes.  These studies identified 266 essential and non-essential GIS genes in which mutations cause increased GCR rates, another 38 candidate GIS genes and 438 cooperating GIS genes that do not suppress GCRs by themselves but act in cooperation with other genes to suppress GCRs.  We have also developed efficient whole genome DNA sequencing (WGS) methods including sequence analysis software for determining the complete structure of rearranged chromosomes.  Our current efforts are focused on: 1) elucidating the mechanisms by which individual genes act to suppress the formation of GCRs, 2) determining the mechanisms that form different types of GCRs, and 3) identifying the complete genetic network that acts to suppress GCRs.
Exploring Genome Instability in Mammalian Cells
One of our long-term goals is to apply insights obtained from S. cerevisiae to the study of genome instability in mammalian cells.  Because genome instability is a characteristic of many human tumors, we have been mining genomics data generated from human tumors.  We have identified 12 different cancers in which there is significant enrichment for defects in human homologues of the S. cerevisiae GIS genes.  These results support the idea that the genome instability seen in many cancers is due to defects in GIS genes.  We are extending these studies in two ways.  First, we are using CRISPR/Cas to knock out the human homologues of the S. cerevisiae GIS genes in human cell lines and test the resulting mutant cell lines for increased genome instability by karyotyping and single cell WGS.  Second, we are using the S. cerevisiae genome instability suppressing gene networks we have discovered to predict candidate therapeutic targets for cancers that have defects in GIS genes.  Thus far, we have been able to identify and validate a new therapeutic target for BRCA1and BRCA2 mutant cancers and are working to identify therapeutic targets for cancers with other types of GIS gene defects.