The coordination between DNA polymerase β and ligase governs the formation of mutagenic repair intermediates as an important determinant of faithful base excision repair

Abstract

Base excision repair (BER) is a critical process for preventing the mutagenic and lethal consequences of DNA damage that arises from endogenous and environmental agents and underlies disease and aging. BER pathway involves a series of sequential enzymatic steps that are tightly coordinated in a process known as the substrate-product channeling between repair proteins so that release of cytotoxic BER intermediates is minimized. DNA ligase I and III catalyze the ultimate ligation step following DNA polymerase (pol) β nucleotide insertion at downstream steps of BER pathway. DNA ligases and DNA polymerases that play roles in DNA replication and repair transactions have long been prime candidates for key regulators of genome stability. However, the ligation efficiency of pol β-promoted mutagenesis products with 3′-DNA mismatches or 8-oxo-2'-deoxyguanosine (8-oxodG) at the downstream steps of BER pathway remains undefined. We show that pol β 8-oxo-2'-deoxyribonucleoside 5'-triphosphate (8-oxodGTP) insertion confounds DNA ligases leading to the formation of ligation failure products with a 5'-adenylate (AMP) block. However, pol β Watson-Crick-like dGTP mismatch insertion opposite T can be efficiently ligated by the BER DNA ligases in vitro. Moreover, we report the mutagenic ligation of pol β 8-oxodGTP insertion products by the LIG1 MgHiFimutant with a perturbed fidelity and enhanced ligation failure by the variants of LIG1 deficiency disease that exhibits the symptoms of developmental abnormalities, immunodeficiency, lymphoma, and predisposition to cancer. Our results also demonstrate that the BER DNA ligases are compromised by subtle changes in all 12 possible noncanonical base pairs at the 3′-end of the nicked repair intermediate. Finally, we found that Aprataxin and Flap Endonuclease 1, as compensatory DNA-end processing enzymes, can remove the 5'-AMP block from the abortive ligation products harboring 3′-8-oxodG or all possible mispairs. These findings contribute to understanding of how the identity of the damaged or mismatch affects the substrate channeling of the repair pathway, importance of mechanism underlying the interplay between pol β and DNA ligases for faithful BER, and how a multi-protein complex can coordinate to maintain the repair efficiency.