Research / Clinical Summary

Richard Kolodner, PhD Professor, Medicine Cancer Genes and Genome Program Contact by Email

Genetic recombination is a central aspect of DNA metabolism, which has a number of essential functions. My laboratory is using Saccharomyces cerevisiae as a model organism to study the mechanism of genetic recombination and related repair events. A related interest is in understanding the molecular mechanisms by which the cell maintains the stability of its genome and prevents the accumulation of mutations and other types of genome rearrangements. Our goal is to reconstitute these reactions in vitro to determine how proteins catalyze them. We are also working on inherited defects in human recombination and repair genes to understand how such defects cause cancer susceptibility and to understand the basic human genetics of these genes. These studies involve the routine use of techniques in bacterial and yeast genetics, protein purification, genomics, mouse genetics and human genetics.

S. cerevisiae is an ideal system for studying eukaryotic recombination and mismatch repair because it is amenable to both genetic and biochemical analysis. Our work on S. cerevisiae recombination is presently focussed on three proteins required for normal levels of meiotic crossing over: (1) the DNA helicase encoded by the MER3 gene; and (2) the MSH4-MSH5 complex whose biochemical function is presently unknown. Our work on DNA mismatch repair is focused on many different genes and the proteins they encode. These include: (1) the three genes which encode RPA, an ssDNA binding protein; (2) PCNA, which is a DNA polymerase accessory factor; (3) DNA polymerase delta; (4) MSH1, a mitochondrial protein, and the MSH2-MSH3 and MSH2-MSH6 nuclear complexes that are mispaired base recognition proteins/complexes required for mismatch repair in the mitochondria and nucleus, respectively; (5) the MLH1-PMS1 and MLH1-MLH3 complexes that interact with the MSH2-MSH3 and MSH2-MSH6 complexes; and (5) EXO1 and RTH1, which are exonucleases that play a role in different types of DNA repair. We are also performing genetic and biochemical experiments to identify additional proteins that function in DNA mismatch repair in order to reconstitute mismatch repair with purified proteins.

Through our studies on DNA repair in S. cerevisiae, we have recently identified a new class of genes that function in preventing the accumulation of deletion mutations and chromosomal translocations that are like the chromosomal rearrangements seen in human cancer cells. These genes fall into at least four classes: (1) genes encoding DNA replication proteins; (2) genes encoding proteins that function in the S-phase checkpoint; (3) genes that encode proteins that are key components of a type of recombination called ”break-induced replication’; and (4) genes that regulate the activity of telomerase. Our future goals are to identify the mechanism of these mutation suppression pathways and to determine the mechanisms by which gross chromosomal rearrangements occur.

Hereditary non-polyposis colon cancer (HNPCC) is a common inherited cancer susceptibility syndrome. We have recently cloned genes encoding the human homologues of the S. cerevisiae mismatch repair genes MSH2 and MLH1 and demonstrated that inherited mutations in these genes are responsible for approximately 50% of HNPCC. In addition, analysis of the MSH6 gene has shown that inherited msh6 mutations cause an attenuated form of inherited cancer susceptibility associated with incomplete penetrance and later age of onset compared with classical HNPCC. As part of these studies, we have developed mice carrying mutations in different mismatch repair genes and are using these mice as a model system to study the development of cancer caused by mismatch repair defects. Our future efforts in this area are focused on trying to identify additional human cancer susceptibility genes that are related to other S. cerevisiae genes we are studying, determining if there is a polygenic form of mismatch repair defective cancer susceptibility, better defining the diseases that mutations in mismatch repair genes cause and to develop chemotherapy directed at mismatch repair defective cancer cells.