Figure 1. Mismatch Repair Pathway.
Overview of Mismatch repair pathway
DNA mismatch repair (MMR) recognizes and repairs erroneous insertion, deletion, and mis-incorporation of bases that can arise during DNA replication and recombination, and repair some forms of DNA damage. It plays an important role in maintaining genomic stability and cellular homeostasis. For example, MMR increases the accuracy of DNA replication by 20–400-fold in Escherichia coli. The mismatch repair machinery distinguishes the newly synthesised strand from the template (parental) to begin the repair process. Nowadays, 2 types of MMR mechanisms have been demonstrated, one is expected to be employed by eukaryotes and the majority of bacteria, and the other is specific to E. coli and some bacteria.
Figure 2. MMR complex
The element of MMR system
Prokaryotic MMR system
The MMR system in E. coli is essentially composed of four identifiable proteins, MutS, MutL, MutH, and UVrs.
MutS is the pivotal protein of the MMR system that can detect mismatches in the double-stranded DNA. It initiates the MMR machinery by recognizing nonspecific interactions. It has two functional domains including a DNA-binding domain and an ATPase/dimerization domain. Both domains of the MutS protein are sterically distant in the inactive state, but conformation changes on recognizing a specific lesion in DNA, ATP or just binding to DNA. Both protein domains affect each other’s function.
MutL protein acts as a mediator between the MutS and other protein complexes in the MMR systems. It creates a link between the MutS mismatch recognization and the excision of the mismatch bases by responsible proteins. MutL interacts with the activated MutS homodimer and sequentially stimulates the endonuclease activity of the MutH protein. In addition, MutL can also function as the recruiter of UvrD, which facilitates its loading onto the DNA lesion site.
MutH is a member of the type II family of restriction endonucleases. It cleaves the mismatch-containing DNA single strand specifically at hemimethylated GATC sites. MutH nicking activity is stimulated by the MMR complex consisting of MutS, MutL, and ATP. The C-terminal helix of MutH protein acts as a molecular attache´ through which MutS and MutL communicate and activate MutH.
UvrD is a DNA helicase II that unwinds DNA starting from the nick generated by MutH. It is loaded onto MutL at the DNA duplex lesion site, which increases its intrinsic helicase activity. The nick generated by MutH serves as a point of entry for single stranded DNA-binding protein and UvrD/helicase II. The loading of UvrD/helicase II at the nick is facilitated through protein–protein interactions with MutL. In addition, many other proteins also take part in this process, including three single strand DNA-specific exonucleases in MMR system, the 3’ -5’ exonucleases ExoI and ExoX and the 5’ -3’ exonucleases, which are responsible for the excision of the newly synthesized DNA strand between the nick and the mismatch.
Figure 3. Methyl-directed mismatch repair
Eukaryotic mismatch repair system
In MMR system of eukaryotes, MSHs play an important role. It is homologous to MutS. Five highly conserved MSHs (MSH2–MSH6) have been found in both yeast and mammals. In the system, two heterodimers of MSHs are found, including MutSa (MSH2/MSH6) and MutSb (MSH2/MSH3). The MutSa complex represents about 80–90% of the eukaryotic cellular level of MSH2. The protein MSH6 of MutSa heterodimer has been found to be responsible for recognizing the mismatch within the DNA duplex. It plays an important role in the recognition of mismatched DNA and in the correction of base–base mispairs as well as lots of insertion/deletion loop (IDL) mispairs. The MutSb heterodimer functions specifically in the repair of IDL mispairs.
In addition, the eukaryotic cell has MLH proteins which are homologs of MutL proteins of E. coli. In total, there are four MutL homologs (MLH1, MLH3, PMS1 and PMS2) in both yeast and mammals. They also play roles by the form of heterodimer. According to previous studied, there are three complexes in eukaryotic cell, such as, MutLa (MLH1–PMS2), MutLb (MLH1–PMS1), and MutLg (MLH1–MLH3). MutLa supports repair initiated by the MutS complex. It is the most active complexes in humans. Little is known about the function of human MutLb complex. MutLg are believed to participate in the repair of a subset of IDLs and also involved in meiotic recombination.
In addition to MSHs and MLHs, eukaryotes also have a wide variety of accessory proteins that are indispensable for the proper functioning of the MMR system, such as, proliferating cell nuclear antigen (PCNA), replication protein A (RPA), and exonuclease I (ExoI). They assist the main MMR proteins in recognizing, binding to, or excising the mismatch from the duplex DNA.
Mismatch repair pathway models
At the mismatch point, the DNA is sharply kinked about 60° towards a narrowed major groove. MutS detects mismatches in the double-stranded DNA by its conserved Phe-X-Glu motif at the N-terminus of bacterial, which initiate the MMR machinery.
The recruitment of related proteins
MutL interacts with the activated homodimer form of MutS and set up a bridge for other protein complexes of MMR. It creates a link between the mismatch recognized by MutS and the proteins responsible for excision of the mismatch, and stimulates the endonuclease activity of the MutH protein.
The formation of a ternary complex
MutS, MutL, and a mismatched DNA formed to the ternary complex.
Excision and gap filling
EXO1 is activated by MutSα and removed the mismatch base, and then RPA will displace the mismatch base. After the excision and replacement of mismatch base are finished, the activity of EXO1 is inhibited by MutSα and MutLα.
Figure 4. Schematic representation of MMR pathway models
Fukui K. DNA mismatch repair in eukaryotes and bacteria. J Nucleic Acids. 2010, 2010. pii: 260512.
Peggy Hsieh and Kazuhiko Yamane. DNA mismatch repair: Molecular mechanism, cancer, and ageing. Mech Ageing Dev. 2008; 129(7-8): 391–407.
Sameer AS, Nissar S, Fatima K. Mismatch repair pathway: molecules, functions, and role in colorectal carcinogenesis. Eur J Cancer Prev. 2014, 23(4):246-57.
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