Junqiu Zhang

Huntington’s disease (HD) is an inherited neurodegenerative disorder caused by an expanded CAG repeat in the huntingtin ( HTT ) gene. The repeat-expanded HTT encodes a mutated HTT (mHTT), which is known to induce DNA double-strand breaks (DSBs), activation of the cGAS–STING pathway, and apoptosis in HD. However, the mechanism by which mHTT triggers these events is unknown. Here, we show that HTT interacts with both exonuclease 1 (Exo1) and MutLα (MLH1–PMS2), a negative regulator of Exo1. While the HTT–Exo1 interaction suppresses the Exo1-catalyzed DNA end resection during DSB repair, the HTT–MutLα interaction functions to stabilize MLH1. However, mHTT displays a significantly reduced interaction with Exo1 or MutLα, thereby losing the ability to regulate Exo1. Thus, cells expressing mHTT exhibit rapid MLH1 degradation and hyperactive DNA excision, which causes severe DNA damage and cytosolic DNA accumulation. This activates the cGAS–STING pathway to mediate apoptosis. Therefore, we have identified unique functions for both HTT and mHTT in modulating DNA repair and the cGAS–STING pathway-mediated apoptosis by interacting with MLH1. Our work elucidates the mechanism by which mHTT causes HD.

Mismatch repair (MMR) is a replication-coupled DNA repair mechanism and plays multiple roles at the replication fork. The well-established MMR functions include correcting misincorporated nucleotides that have escaped the proofreading activity of DNA polymerases, recognizing nonmismatched DNA adducts, and triggering a DNA damage response. In an attempt to determine whether MMR regulates replication progression in cells expressing an ultramutable DNA polymerase ɛ (Polɛ), carrying a proline-to-arginine substitution at amino acid 286 (Polɛ-P286R), we identified an unusual MMR function in response to hydroxyurea (HU)-induced replication stress. Polɛ-P286R cells treated with hydroxyurea exhibit increased MRE11-catalyzed nascent strand degradation. This degradation by MRE11 depends on the mismatch recognition protein MutSα and its binding to stalled replication forks. Increased MutSα binding at replication forks is also associated with decreased loading of replication fork protection factors FANCD2 and BRCA1, suggesting blockage of these fork protection factors from loading to replication forks by MutSα. We find that the MutSα-dependent MRE11-catalyzed fork degradation induces DNA breaks and various chromosome abnormalities. Therefore, unlike the well-known MMR functions of ensuring replication fidelity, the newly identified MMR activity of promoting genome instability may also play a role in cancer avoidance by eliminating rogue cells.

Cancer is instigated by mutator phenotypes, including deficient mismatch repair and p53-associated chromosomal instability. More recently, a distinct class of cancers was identified with unusually high mutational loads due to heterozygous amino acid substitutions (most commonly P286R) in the proofreading domain of DNA polymerase ε, the leading strand replicase encoded by POLE . Immunotherapy has revolutionized cancer treatment, but new model systems are needed to recapitulate high mutational burdens characterizing human cancers and permit study of mechanisms underlying clinical responses. Here, we show that activation of a conditional LSL-Pole ^P286R allele in endometrium is sufficient to elicit in all animals endometrial cancers closely resembling their human counterparts, including very high mutational burden. Diverse investigations uncovered potentially novel aspects of Pole -driven tumorigenesis, including secondary p53 mutations associated with tetraploidy, and cooperation with defective mismatch repair through inactivation of Msh2 . Most significantly, there were robust antitumor immune responses with increased T cell infiltrates, accelerated tumor growth following T cell depletion, and unfailing clinical regression following immune checkpoint therapy. This model predicts that human POLE -driven cancers will prove consistently responsive to immune checkpoint blockade. Furthermore, this is a robust and efficient approach to recapitulate in mice the high mutational burdens and immune responses characterizing human cancers.