The observation in this article is intriguing in the context of eukaryotic mismatch repair. Transient hemimethylated CpG sites is possible in eukaryotes during active replication. It would be interesting to further explore if the binding of MutSa on hemimethylated CpG has any role in further activation of mismatch repair, or on the recruitment of any other partner proteins for that matter. It would also be interesting to study if the presence of more than one hemimethylated CpG sites on a single oligo increases the binding affinity or stoichiometry. However, that would have to be investigated in a future study. As an individual observation this is very relevant, and the gel mobility shift assays presented in this work unambiguously indicates significant binding of MutSa with hemimethylated DNA.
This reviewer has some questions regarding the data provided:
1) Figure 1D (Quantification of EMSA), the y-axis is denoted as “% of bound complex/free oligo”. The unit is not clear. The y-axis values seem to be in fraction rather than a percentage. Or, are the authors suggesting the intensity of signal in the complex formed of 1000 ng of MutSa with hemimethylated DNA substrate is approximately 1% of the free oligo seen in the gel?
Additionally, please include the quantification for the T/G mismatch substrate also in this graph.