GEOapplication/xmlftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE319nnn/GSE319524/primaryOK200Other Homo sapiensSaccharomyces cerevisiaeOtherhttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE319524GEOGSEfalseTranscription-coupled repair removes cytotoxic DNA methylation damage and promotes mutational strand asymmetry in gliomasTranscription-coupled nucleotide excision repair (TC-NER) is known to repair bulky DNA adducts such as UV damage. However, whether TC-NER repairs small base damage is less understood. Alkylating agents induce cytotoxic base modifications (e.g., methylation) and are widely used in chemotherapy. Here we used a genome-wide methylation damage mapping method, N-methylpurine sequencing 2.0 (NMP-seq 2.0), and generated high-resolution methylation damage and repair profiles in human cells treated by methyl methanesulfonate (MMS). Our data revealed a repair strand asymmetry in base excision repair (BER) deficient cells lacking the N-methylpurine DNA glycosylase (MPG, also known as AAG), with higher repair rate on the transcribed strand (TS) relative to the non-transcribed strand (NTS). MMS induces N7-methylguanine (7meG) and N3-methyladenine (3meA). Interestingly, the repair asymmetry was only found for 3meA, but not for 7meG, which is less cytotoxic and mutagenic than 3meA. Further knockout of TC-NER genes, CSB or UVSSA, significantly sensitized AAG-deficient cells to MMS and abolished the repair asymmetry, indicating that the TS-biased repair of 3meA is dependent on TC-NER. The methylating agent temozolomide (TMZ) is used for glioma treatment. By analyzing glioma mutation data, we found that TMZ-treated gliomas accumulated fewer G>A mutations on the TS relative to the NTS, a phenomenon that can be attributed to strand-biased repair of O6-methylguanine (O6meG) by TC-NER. Finally, we show that knockout of the TC-NER factor CSB significantly sensitized glioma cells to TMZ. These findings indicate that TC-NER repairs cytotoxic methylation damage (e.g., 3meA and O6meG) on the TS, promotes mutational strand asymmetry in TMZ-treated gliomas, and maintains resistance to alkylating agents.2026/04/30GSE319524GSM9519076GSM9519098GSM9519077GSM9519074GSM9519096GSM9519097GSM9519075GSM9519078GSM9519079GSM9519090GSM9519091GSM9519094GSM9519073GSM9519095GSM9519092GSM9519093GSM9519087GSM9519088GSM9519085GSM9519086GSM9519089GSM9519080GSM9519083GSM9519084GSM9519081GSM95190822467618249319524 Homo sapiensSaccharomyces cerevisiae