Phosphorylated and unphosphorylated forms of human single-stranded DNA-binding protein are equally active in simian virus 40 DNA replication and in nucleotide excision repair.
The trimeric human single-stranded DNA-binding protein (HSSB; also called RP-A) plays an essential role in DNA replication, nucleotide excision repair, and homologous DNA recombination. The p34 subunit of HSSB is phosphorylated at the G1/S boundary of the cell cycle or upon exposure of cells to DNA damage-inducing agents including ionizing and UV radiation. We have previously shown that the phosphorylation of p34 is catalyzed by both cyclin-dependent kinase-cyclin A complex and DNA-dependent protein kinase. In this study, we investigated the effect of phosphorylation of p34 by these kinases on the replication and repair function of HSSB. We observed no significant difference with the unphosphorylated and phosphorylated forms of HSSB in the simian virus 40 DNA replication or nucleotide excision repair systems reconstituted with purified proteins. The phosphorylation status of the p34 subunit of HSSB was unchanged during the reactions. We suggest that the phosphorylated HSSB has no direct effect on the basic mechanism of DNA replication and nucleotide excision repair reactions in vitro, although we cannot exclude a role of p34 phosphorylation in modulating HSSB function in vivo through a yet poorly understood control pathway in the cellular response to DNA damage and replication.
Pan
Z Q
ZQ
Graduate Program in Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Park
C H
CH
Amin
A A
AA
Hurwitz
J
J
Sancar
A
A
D004260:DNA Repair; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D010766:Phosphorylation; D013539:Simian virus 40 / Q000235:genetics / Q000378:metabolism*
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances