AP endonuclease and poly(ADP-ribose) polymerase-1 interact with the same base excision repair intermediate

Base excision repair (BER) is a defense system that protects cells from deleterious effects secondary to modified or missing DNA bases. BER is known to involve apurinic/apyrimidinic endonuclease (APE) and DNA polymerase B (B-pol) among other enzymes, and recent studies have suggested that poly(ADP-ribose) polymerase- 1 (PARP-1) also plays a role by virtue of its binding to BER intermediates. The main role of APE is cleavage of the DNA backbone at abasic sites, and the enzyme also can catalyze 3'- to 5'-exonuclease activity at the cleaved abasic site. Photocross-linking studies with mouse embryonic fibroblast (MEF) cell extracts described here indicated that APE and PARP- 1 interact with the same APE-cleaved abasic site BER intermediate. The model BER intermediate used includes a synthetic abasic site sugar, i.e. tetrahydrofuran (THF), in place of the natural deoxyribose. APE cross-linked efficiently with this intermediate, but not with a molecule lacking the 5'-THF phosphate group, and the same property was demonstrated for PARP- 1. The addition of purified APE to the MEF extract reduced the amount of PARP- I cross-linked to the BER intermediate, suggesting that APE can compete with PARP- 1. APE and PARP- 1 were antagonists of each other in in vitro BER related reactions on this model BER intermediate. These results suggest that PARP- I and APE can interact with the same BER intermediate and that competition between these two proteins may influence their respective BER related functions. (C) 2004 Elsevier B.V. All rights reserved.