Analysis of interactions between mismatch repair initiation factors and the replication processivity factor PCNA

J Mol Biol. 2006 Jan 13;355(2):175-84. doi: 10.1016/j.jmb.2005.10.059. Epub 2005 Nov 8.

Abstract

In eukaryotes, the DNA replication factor PCNA is loaded onto primer-template junctions to act as a processivity factor for DNA polymerases. Genetic and biochemical studies suggest that PCNA also functions in early steps in mismatch repair (MMR) to facilitate the repair of misincorporation errors generated during DNA replication. These studies have shown that PCNA interacts directly with several MMR components, including MSH3, MSH6, MLH1, and EXO1. At present, little is known about how these interactions contribute to the mismatch repair mechanism. The interaction between MLH1 and PCNA is of particular interest because MLH1-PMS1 is thought to act as a matchmaker to signal mismatch recognition to downstream repair events; in addition, PCNA has been hypothesized to act in strand discrimination steps in MMR. Here, we utilized both genetic and surface plasmon resonance techniques to characterize the MLH1-PMS1-PCNA interaction. These analyses enabled us to determine the stability of the complex (K(D) = 300 nM) and to identify residues (572-579) in MLH1 and PCNA (126,128) that appear important to maintain this stability. We favor a model in which PCNA acts as a scaffold for consecutive protein-protein interactions that allow for the coordination of MMR steps.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Animals
  • Base Pair Mismatch*
  • Blotting, Far-Western
  • Carrier Proteins / metabolism
  • DNA Repair*
  • DNA, Fungal / metabolism
  • DNA-Binding Proteins / metabolism
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism*
  • Humans
  • Mice
  • Molecular Sequence Data
  • MutL Protein Homolog 1
  • MutL Proteins
  • MutS Homolog 2 Protein / metabolism
  • Mutation
  • Proliferating Cell Nuclear Antigen / metabolism*
  • Protein Structure, Tertiary
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Surface Plasmon Resonance

Substances

  • Adaptor Proteins, Signal Transducing
  • Carrier Proteins
  • DNA, Fungal
  • DNA-Binding Proteins
  • Fungal Proteins
  • MLH1 protein, S cerevisiae
  • MSH6 protein, S cerevisiae
  • PMS1 protein, S cerevisiae
  • Proliferating Cell Nuclear Antigen
  • Saccharomyces cerevisiae Proteins
  • MSH2 protein, S cerevisiae
  • MutL Protein Homolog 1
  • MutL Proteins
  • MutS Homolog 2 Protein