Human pluripotent stem cells (PSCs) are presumed to have robust DNA repair pathways to ensure genome stability. PSCs likely need to protect against mutations that would otherwise be propagated throughout all tissues of the developing embryo. How these cells respond to genotoxic stress has only recently begun to be investigated. While PSCs appear to respond to certain forms of damage more efficiently than somatic cells, some DNA damage response pathways such as the replication stress response may be lacking. Not all DNA repair pathways have been well characterized in PSCs to date, including the DNA mismatch repair (MMR) pathway. MMR maintains genomic stability by repairing DNA polymerase errors. MMR is also involved in the induction of cell cycle arrest and apoptosis in response to certain exogenous DNA damaging agents. Here, we examined MMR function in PSCs. We have demonstrated that PSCs contain a robust MMR pathway and are highly sensitive to DNA alkylation damage in a MMR-dependent manner. Interestingly, the nature of this alkylation response differs from that previously reported in somatic cell types. In somatic cells, a permanent G2/M cell cycle arrest is induced in the second cell cycle after DNA damage. The PSCs, however, directly undergo apoptosis in the first cell cycle. This response reveals that PSCs rely on apoptotic cell death as an important defense to avoid mutation accumulation. Our results also suggest an alternative molecular mechanism by which the MMR pathway can induce a response to DNA damage that may have implications for tumorigenesis.