The role of METTL3 and ADP-ribosylation in DNA repair

González Leal, Claudia Patricia (2025): The role of METTL3 and ADP-ribosylation in DNA repair. Dissertation, LMU München: Medizinische Fakultät

Abstract

The DNA damage response (DDR) is a critical cellular mechanism that maintains genomic integrity. Failure of the DDR can lead to cell cycle arrest, cell death, or the emer-gence of diseases such as cancer. A key player in DDR is the Poly(ADP-ribose) polymerase 1 (PARP1), which detects DNA lesions and triggers a rapid burst of ADP-ribosylation (PARylation). This post-translational modification facilitates various cellular processes, including chromatin decompaction, recruitment of DDR proteins, modulation of their activity, and scaffolding of DDR assemblies. Growing evidence indicates a role for RNA and RNA-binding proteins in the DDR, with RNA serving as a template for RNA-mediated repair, stalled RNA polymerase serving as a surveillance mechanism of DNA integrity, and RNA-binding proteins that play multiple roles in DDR pathways. Recent studies have highlighted the overlap between RNA-binding proteins and PAR interactors, suggesting a complex interplay between PARylation and RNA biology. This study focuses on METTL3/14, a complex responsible for catalyzing N6-methyladenosine (m6A) methylation of RNA, which has been shown to recruit to DNA damage sites in a PARP-dependent manner. In this study, we characterized the recruitment of METTL3/14 in real-time using live-cell imaging coupled with laser microirradiation. We confirmed that METTL3/14 rapidly recruited to DNA damage sites, with its recruitment being dependent on active PARylation. Our assays suggest that the methyltransferases recruit independently of each other, that the C-terminal region of METTL3, which contains the methyltransferase domain, is sufficient for recruitment, and that RNA contributes to the accumulation of the complex at DNA lesions. Furthermore, we provide evidence that METTL3/14 can bind to PAR and RNA in vitro and that high concentrations of PAR can displace, although not out-compete RNA. Moreover, METTL3 deficient cells exposed to UV exhibited delayed 6-4PP and CPD repair, delayed transcription restart, and ATF3 accumulation. Moreover, these cells are sensitive to Illudin S, suggesting a role for METTL3 in transcription-coupled nucleotide excision repair (TC-NER). Additionally, we explored the therapeutic potential of combining METTL3 and PARP inhibitors, which showed additive antiproliferative effects in leukemia and breast cancer models. Our findings elucidate the complex interplay between DNA repair, RNA biology, and PARylation, offering insights into METTL3/14's roles and potential clinical applications.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: METTL3, poly(ADP-ribosylation), PAR, m6A, DNA damage
• Themengebiete: 600 Technik, Medizin, angewandte Wissenschaften ; 600 Technik, Medizin, angewandte Wissenschaften > 610 Medizin und Gesundheit
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 12. Dezember 2025
• 1. Bericht­erstatter:in: Ladurner, Andreas G.