Impact of polβ/XRCC1 Interaction Variants on the Efficiency of Nick Sealing by DNA Ligase IIIα in the Base Excision Repair Pathway

• Published in: Journal of molecular biology 2024-02, Vol.436 (4), p.168410-168410, Article 168410
• Main Authors: Almohdar, Danah, Gulkis, Mitchell, Ortiz, Abigail, Tang, Qun, Sobol, Robert W., Çağlayan, Melike
• Format: Article
• Language: English
• Subjects: base excision repair; DNA Breaks, Single-Stranded; DNA Ligase ATP - chemistry; DNA ligase IIIα; DNA Polymerase beta - chemistry; DNA polymerase β; Excision Repair; Humans; Protein Binding; repair pathway coordination; X-ray cross-complementing protein 1; X-ray Repair Cross Complementing Protein 1 - chemistry; X-ray Repair Cross Complementing Protein 1 - genetics
• ISSN: 0022-2836; 1089-8638; 1089-8638
• DOI: 10.1016/j.jmb.2023.168410

[Display omitted] •Base excision repair requires coordination between polβ and DNA ligase and is scaffolded by XRCC1.•Polβ colon cancer associated variant T304 exhibits a reduced interaction with XRCC1.•Polβ mutants regulating diminished ubiquitin-mediated degradation do not form repair complex with XRCC1.•Polβ variants that cannot interact with XRCC1 leads to ligation failure by DNA ligase IIIα.•Polβ/XRCC1/DNA ligase IIIα interactome is required for accurate BER pathway coordination. Base excision repair (BER) requires a coordination from gap filling by DNA polymerase (pol) β to subsequent nick sealing by DNA ligase (LIG) IIIα at downstream steps of the repair pathway. X-ray cross-complementing protein 1 (XRCC1), a non-enzymatic scaffolding protein, forms repair complexes with polβ and LIGIIIα. Yet, the impact of the polβ mutations that affect XRCC1 interaction and protein stability on the repair pathway coordination during nick sealing by LIGIIIα remains unknown. Our results show that the polβ colon cancer-associated variant T304 exhibits a reduced interaction with XRCC1 and the mutations in the interaction interface of V303 loop (L301R/V303R/V306R) and at the lysine residues (K206A/K244A) that prevent ubiquitin-mediated degradation of the protein exhibit a diminished repair protein complex formation with XRCC1. Furthermore, we demonstrate no significant effect on gap and nick DNA binding affinity of wild-type polβ by these mutations. Finally, our results reveal that XRCC1 leads to an efficient channeling of nick repair products after nucleotide incorporation by polβ variants to LIGIIIα, which is compromised by the L301R/V303R/V306R and K206A/K244A mutations. Overall, our findings provide insight into how the mutations in the polβ/XRCC1 interface and the regions affecting protein stability could dictate accurate BER pathway coordination at the downstream steps involving nick sealing by LIGIIIα.