Vanhuele, S. | Search bioRxiv

bioRxiv2026

Human PMS1-dependent non-canonical mismatch repair converges with MBD4 to repair 5-methylcytosine deamination

CpG dinucleotides are hotspots for mutagenesis by spontaneous deamination of 5-methylcytosine (5mC) into thymine, resulting in T:G mismatches that can lead to C>T transitions. These mutations are a hallmark of aging and cancer and a major force shaping the evolution of vertebrate genomes. We have previously uncovered MBD4 as the primary base excision repair (BER) glycosylase responsible for 5mC deamination repair. In this study, we employ a cytosine base-editing system, comprising an APOBEC1 deaminase fused to a catalytically dead Cas9, to induce targeted cytosine deamination in native chromatin and track its repair. This approach reveals that MBD4 cooperates with a non-canonical branch of mismatch repair (MMR) to elicit 5mC deamination repair. We demonstrate that MBD4 activity depends on MMR complexes MutS (MSH2-MSH6) and MutL{beta} (MLH1-PMS1), but not on post-replicative MMR elicited by the MutL (MLH1-PMS2) complex. We find that PMS1 loss phenocopies the genome-wide CpG>TpG hypermutated profile associated with MBD4 deficiency, uncovering that 5mC deamination repair may represent one of the primary functions of the MutL{beta} complex. The mutational landscape of MMR-deficient tumors aligns with our experimental results, showing that replication-independent CpG>TpG mutagenesis partly contributes to the mutational burden of tumors inactivated for MLH1, MSH2 and MSH6. Finally, using structural predictions alongside biochemical validation, we show that MBD4 physically interacts with MutL{beta} in an MLH1-dependent manner, illuminating the structural basis for the convergence of the BER and MMR pathways. Altogether, we uncover a novel function of non-canonical MMR that underscores its interplay with BER in safeguarding genomic integrity against damage to methylated DNA.

molecular biology↗

bioRxiv2022

Optical Genome Mapping for detecting Homologous Recombination Deficiency (HRD) in human breast cancers

Homologous recombination deficiency (HRD) leads to genomic instability that marks HRD tumor genome with a specific genomic scar. Present in many cancers, HRD is important to be detected as it is associated with a hyper-sensitivity to some classes of drugs, in particular the PARP inhibitors. Here, we investigated the use of structural variants (SVs) detected by the Optical Genome Mapping (OGM) technology as biomarkers to identify HRD tumors. We analyzed SVs data obtained by OGM from 37 samples of triple-negative breast cancer or high grade ovarian cancer with the known HRD status. We found that HRD cases were enriched with duplications and reciprocal translocations, while nonHRD cases were enriched with inversions. The number of translocations, defined as inter-chromosomal or intra-chromosomal rearrangements of more 5Mb were similar in HRD and nonHRD cases. We defined isolated translocations as the subset of translocations having no other translocation within 2 megabase zone around both junctions, and demonstrated that the number of isolated translocations perfectly discriminated HRD and nonHRD cases in the training series. Validation series consisting from 26 cases showed 20% false positive and zero false negative error rate, which proved isolated translocations to be 100% sensitive and 80% specific SV marker of HRD. Our results demonstrate that the OGM technology is an affordable way of getting an insight of the structural variants present in solid tumors, even with low tumoral cellularity. It represents a promising technology for HRD diagnosis, where a single marker already gives 80% correct recognition.

cancer biology↗