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  • HMCES Maintains Replication Fork Progression and Prevents Double-Strand Breaks in Response to APOBEC Deamination and Abasic Site Formation.

HMCES Maintains Replication Fork Progression and Prevents Double-Strand Breaks in Response to APOBEC Deamination and Abasic Site Formation.

Cell reports (2020-06-04)
Kavi P M Mehta, Courtney A Lovejoy, Runxiang Zhao, Darren R Heintzman, David Cortez
ABSTRACT

5-Hydroxymethylcytosine (5hmC) binding, ES-cell-specific (HMCES) crosslinks to apurinic or apyrimidinic (AP, abasic) sites in single-strand DNA (ssDNA). To determine whether HMCES responds to the ssDNA abasic site in cells, we exploited the activity of apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3A (APOBEC3A). APOBEC3A preferentially deaminates cytosines to uracils in ssDNA, which are then converted to abasic sites by uracil DNA glycosylase. We find that HMCES-deficient cells are hypersensitive to nuclear APOBEC3A localization. HMCES relocalizes to chromatin in response to nuclear APOBEC3A and protects abasic sites from processing into double-strand breaks (DSBs). Abasic sites induced by APOBEC3A slow both leading and lagging strand synthesis, and HMCES prevents further slowing of the replication fork by translesion synthesis (TLS) polymerases zeta (Polζ) and kappa (Polκ). Thus, our study provides direct evidence that HMCES responds to ssDNA abasic sites in cells to prevent DNA cleavage and balance the engagement of TLS polymerases.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
MISSION® esiRNA, targeting human APEX2
Sigma-Aldrich
5-Chloro-2′-deoxyuridine, thymidine analog
Sigma-Aldrich
JH-RE-06, ≥98% (HPLC)
Sigma-Aldrich
MISSION® esiRNA, targeting human APEX1
Sigma-Aldrich
Anti-HMCES antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution
Sigma-Aldrich
Anti-APOBEC3A antibody produced in rabbit, affinity isolated antibody