Self-reversal facilitates the resolution of HMCES DNA-protein crosslinks in cells

• Published in: Cell reports (Cambridge) Vol. 42; no. 11; p. 113427
• Main Authors: Rua-Fernandez, Jorge, Lovejoy, Courtney A., Mehta, Kavi P.M., Paulin, Katherine A., Toudji, Yasmine T., Giansanti, Celeste, Eichman, Brandt F., Cortez, David
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 28-11-2023; Elsevier
• Subjects: abasic site; base excision repair; CP: Molecular biology; DNA - genetics; DNA Damage; DNA Repair; DNA Replication; DNA, Single-Stranded; Proteins - genetics; replication stress; SPRTN; SRAP; replication stress; DNA damage; base excision repair; SRAP; SPRTN; CP: Molecular biology; abasic site
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2023.113427

Abasic sites are common DNA lesions stalling polymerases and threatening genome stability. When located in single-stranded DNA (ssDNA), they are shielded from aberrant processing by 5-hydroxymethyl cytosine, embryonic stem cell (ESC)-specific (HMCES) via a DNA-protein crosslink (DPC) that prevents double-strand breaks. Nevertheless, HMCES-DPCs must be removed to complete DNA repair. Here, we find that DNA polymerase α inhibition generates ssDNA abasic sites and HMCES-DPCs. These DPCs are resolved with a half-life of approximately 1.5 h. HMCES can catalyze its own DPC self-reversal reaction, which is dependent on glutamate 127 and is favored when the ssDNA is converted to duplex DNA. When the self-reversal mechanism is inactivated in cells, HMCES-DPC removal is delayed, cell proliferation is slowed, and cells become hypersensitive to DNA damage agents that increase AP (apurinic/apyrimidinic) site formation. In these circumstances, proteolysis may become an important mechanism of HMCES-DPC resolution. Thus, HMCES-DPC formation followed by self-reversal is an important mechanism for ssDNA AP site management.
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•ssDNA generation by Polα inhibition generates abasic sites protected by an HMCES-DPC•HMCES-DPC self-reversal is favored by duplex DNA formation and requires the residue E127•Mutation of E127 delays HMCES-DPC resolution in cells and reduces cell proliferation
HMCES forms a reversible DNA-protein crosslink (DPC) to abasic sites in single-stranded DNA to prevent cleavage and double-strand break formation. Rua-Fernandez et al. demonstrate that an HMCES DPC self-reversal reaction is important to resolve the crosslink in cells. Inactivating self-reversal reduces cell fitness.