TY - JOUR
T1 - Requirement for PCNA and RPA in interstrand crosslink-induced DNA synthesis
AU - Li, Lei
AU - Peterson, Carolyn A.
AU - Zhang, Xiaoshan
AU - Legerski, Randy J.
N1 - Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2000/3/15
Y1 - 2000/3/15
N2 - Proliferating nuclear cell antigen (PCNA) and replication protein A (RPA) have proven to be essential elements in many aspects of DNA metabolism including replication, repair and recombination. We have developed an in vitro assay in which the presence of an interstrand crosslink stimulates the incorporation of radiolabeled nucleotides into both damaged and undamaged plasmid DNAs. Using this assay we have investigated the roles of PCNA and RPA in crosslink-induced DNA synthesis. p21, a potent inhibitor of PCNA, was found to strongly inhibit crosslink-induced incorporation. Addition of exogenous PCNA partially restored the resynthesis activity. Likewise, neutralization of RPA by monoclonal antibodies also inhibited incorporation, but the effect was somewhat more pronounced on the undamaged plasmid than the damaged plasmid. Addition of excess RPA also partially reversed antibody inhibition. These results indicate that both PCNA and RPA are required for efficient in vitro DNA resynthesis induced by interstrand crosslinks.
AB - Proliferating nuclear cell antigen (PCNA) and replication protein A (RPA) have proven to be essential elements in many aspects of DNA metabolism including replication, repair and recombination. We have developed an in vitro assay in which the presence of an interstrand crosslink stimulates the incorporation of radiolabeled nucleotides into both damaged and undamaged plasmid DNAs. Using this assay we have investigated the roles of PCNA and RPA in crosslink-induced DNA synthesis. p21, a potent inhibitor of PCNA, was found to strongly inhibit crosslink-induced incorporation. Addition of exogenous PCNA partially restored the resynthesis activity. Likewise, neutralization of RPA by monoclonal antibodies also inhibited incorporation, but the effect was somewhat more pronounced on the undamaged plasmid than the damaged plasmid. Addition of excess RPA also partially reversed antibody inhibition. These results indicate that both PCNA and RPA are required for efficient in vitro DNA resynthesis induced by interstrand crosslinks.
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U2 - 10.1093/nar/28.6.1424
DO - 10.1093/nar/28.6.1424
M3 - Article
C2 - 10684938
AN - SCOPUS:0034653777
SN - 0305-1048
VL - 28
SP - 1424
EP - 1427
JO - Nucleic acids research
JF - Nucleic acids research
IS - 6
ER -