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MSH2 and Human Nonpolyposis Colon Cancer

MSH2 and Human Nonpolyposis Colon Cancer. Yael Aschner. Presentation Outline. What is MMR? What is MSH2? What is HNPCC?. DNA Mismatch Repair (MMR). Identified in bacteria Mechanism for fixing errors in DNA replication

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MSH2 and Human Nonpolyposis Colon Cancer

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  1. MSH2 and Human Nonpolyposis Colon Cancer Yael Aschner

  2. Presentation Outline • What is MMR? • What is MSH2? • What is HNPCC?

  3. DNA Mismatch Repair (MMR) • Identified in bacteria • Mechanism for fixing errors in DNA replication • The frequency of mutations introduced by DNA polymerase during DNA replication is 1/1012 base pairs; of these, 99.9% are repaired by MMR.

  4. MMR in E. coli Three proteins • MutS • MutL • MutH

  5. MMR in E. coli • MutS performs first step of MMR: recognition of base/base mismatches and small insertion or deletion loops (IDL) • ATPase acting as an asymmetric homodimer • Translocates along DNA and promotes DNA loop formation

  6. MMR in E. coli • MutS undergoes conformation change and interacts with MutL in the presence of ATP • MutL stimulates MutH endonuclease activity, which removes newly replicated DNA fragment with mismatch • DNA polymerase fills in the gap

  7. MMR in E. coli

  8. MutS Homolog 2 (MSH2) • Human homolog of E. coli MutS • Chromosme 2p • 15 exons • Encodes a 934 amino acid protein • Involved in the recognition of mismatched bases and the initiation of the MMR reaction

  9. E. coli MutS Crystal Structure The MutS monomer (green) recognizes the G:T mismatch in the DNA (red) and has ADP bound (at the bottom of the figure in red). The monomer in blue provides the non-specific DNA binding domains and an empty ATPase domain

  10. Eukaryotic MMR • Highly conserved • No MutH homolog • Six homologs of MutS (MSH1-MSH6) • Three are needed for initiation of the repair process: MSH2, MSH3, MSH6 • MSH2 forms a heteroduplex with MSH6 or MSH3, forming MutSa and MutSb, respectively

  11. MutSa • MutSa can bind to most base-base mismatches and loops of one or a few nucleotides • Present at higher concentrations • Primary mismatch recognition factor • Responsible for most mismatch recognition

  12. MutSb • MutSb can bind to IDLs with up to ~8 unpaired nucleotides • Not involved in base-base mismatches • Backup system • Some overlap between the complexes • MSH2 acts as a scaffold while MSH3 and MSH6 provide specific binding

  13. Mismatch Recognition

  14. MSH2 Inactivation • Inability to repair base-base mismatches and small nucleotide insertions and deletions that occur as the result of misalignment or erroneous DNA replication • Acts as a TS gene  CANCER

  15. Human Nonpolyposis Colon Cancer (HNPCC) • Associated with deficiency in MMR • Syndrome characterized by familial predisposition to colorectal carcinoma and extracolonic cancers • Gastrointestinal, urological, female reproductive tracts • Most common hereditary colorectal cancer syndrome • 2%-5% of all colon cancers in Western population • Very high penetrance (~85%) • Early onset • 42 years • Autosomal dominant inheritance (cellular level recessive) • Treatment: removal of all or most of the colon

  16. Colon Adenoma of the Large Intestine • Precursor polyps (or adenomas) appears to be no greater than the number found in the general population • These polyps are thought to progress more rapidly to cancer and form at earlier ages

  17. How is MSH2 Mutated in HNPCC? • 40% of HNPCC mutations are in MSH2 gene • Predominantly small deletions, insertions, splice site alterations, nonsense or missense mutations • MSH2 is the most common gene to have deletions • Range from one exon to whole gene • MSH2 or MLH1 mutations responsible for 50%-80% of HNPCC families • 20% of sporadic colorectal cancers due to MMR mutations

  18. HNPCC Inheritance • Individuals heterozygous for mutations • Two Hit Hypothesis • Functional wild type allele lost by somatic mutation event • MMR inactivated • Accumulation of mutations

  19. Loss of Heterozygosity

  20. Mouse Models for HNPCC(Wei, et al) • Msh2+/- mutants indistinguishable from wild type • Homozygous Msh2-/- mice all died by 12 months (50% by 6 months) • Strong predisposition for tumor phenotype • Cell extracts lost ability to repair single-base mispairs and IDLs of up to four nucleotides • Tissues displayed high mutation frequencies • Small intestinal tumors, lymphoid tumors

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