Tumors belonging to the MSI subtype displayed strikingly different associations between epigenetic features and mutation patterns. We observed little association between mutation rate and open chromatin marks or replication timing in MSI tumors. This is likely because mismatch repair (MMR) deficient MSI tumors have lost MMR-coupled enhanced repair efficiency at early replicating open-chromatin regions31. In addition, we found that MSI mutation profiles showed a strong positive association with heterochromatin (H3K9me3) and repressive domains (H3K27me3) (Fig. 1b). This is in contrast with a previous study by Supek et al. reporting that mutations generated after MMR inactivation are no longer enriched in heterochromatin regions, arguing that genome-wide regional mutation rate variation is mostly a result of MMR31. Instead, our data suggests that, in addition to MMR, other repair or mutational processes may further contribute to variation of the GC mutation landscape. Principal component analysis (PCA) on the correlation matrix between the mutation profiles of individual tumors and the epigenetic covariates also revealed MSI tumors as a distinct cluster (Fig. 1c). Accordingly, we removed the small number of MSI tumors (N = 19) from the discovery cohort to ensure all tumors had similar mutational biases.