Biology Reference
In-Depth Information
MMR has been well characterized in bacteria, yeast, and mammalian cells. In
mammalian cells, the MMR pathway involves two heterodimers called MutS and
MutL. MutS heterodimers contain MSH2 complexed with either MSH6 or
MSH3. MutL heterodimers contain MLH1 complexed with either PMS2 or
MLH3. In addition to these two complexes, a number of other proteins are
required including RFC, EXO1, RPA, and PCNA. MutS heterodimers are re-
sponsible for ''sensing'' or recognizing the mismatch followed by recruitment of
the MutL heterodimer to the chromatin.
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The two heterodimers then bind ATP
and undergo a conformational change, resulting in a protein that can more freely
diffuse along the DNA in either direction. This suggests that the MutS-MutL
complex may be functioning as a ''sliding clamp'' on the DNA. The MutS-MutL
complex diffuses across the region containing the DNA mismatch and meets
either RPA at the 5
0
terminus or PCNA at the 3
0
terminus. After this, it loads
and activates EXO1 to degrade the faulty strand until the mismatch is removed.
Finally, DNA polymerase
d
fills the gap and DNA ligase I completes the process.
MMR is strongly associated with replication, a process that induces signif-
icant chromatin remodeling and has a number of associated histone modifica-
tion events.
88-90
However, it is not yet fully defined as to whether MMR
actually requires any specific chromatin remodeling or any histone modifica-
tions. Most biochemical studies have been carried out on naked DNA sub-
strates so the effect of chromatin on this process is not well studied.
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However,
studies using the yeast genome have demonstrated that MMR has different
efficiencies in different regions, which is suggestive of chromosome structure
being an influence on MMR.
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DNAmodifications resulting from treatment with alkylating agents are known
to activateMMR, but there are subtle differences depending on the dosage. High
doses lead to a G2 cell cycle arrest after one round of replication and PCNA is no
longer present in the chromatin-boundMutS-MutL complex, but with low doses,
the same effects are seen only after the second round of replication.
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This latter
effect might be a method of maintaining the MMR complexes on the chromatin
longer such that other proteins can be recruited and activated.
Although aberrant DNA methylation has been shown to be responsible for
the silencing of MMR genes, studies on hypoxia and stem cells have suggested
that epigenetic chromatin inactivation is responsible for silencing the MMR
genes MLH1 and MSH6.
93
Chromatin immunoprecipitation identified hypoa-
cetylated/hypermethylated histone H3 lysine 9 (H3K9me3) as being present,
which was associated with impaired SP1 binding to the promoters of these
genes. Treatment with HDAC inhibitors increased histone H3K9 acetylation,
which in turn increased SP1 activity on the promoters, and hence increased
expression of MLH1 and MSH6. Therefore, a deregulated MMR pathway may
result in genomic instability in stem cells, thus leading to their malignant
transformation into cancer stem cells.
