Activation of SV40 DNA replication in vitro by cellular protein phosphatase 2A.
We have made use of the cell-free SV40 DNA replication system to identify and characterize cellular proteins required for efficient DNA synthesis. One such protein, replication protein C (RP-C), was shown to be involved with SV40 large T antigen in the early stages of viral DNA replication in vitro. We demonstrate here that RP-C is identical to the catalytic subunit of cellular protein phosphatase 2A (PP2Ac). The purified protein dephosphorylates specific phosphoamino acid residues in T antigen, consistent with the hypothesis that SV40 DNA replication is regulated by modulating the phosphorylation state of the viral initiator protein. We also show that purified RP-C/PP2Ac preferentially stimulates SV40 DNA replication in extracts from early G1 phase cells. This finding suggests that the activity of a cellular factor that influences the net phosphorylation state of T antigen is cell cycle dependent.
Virshup
D M
DM
Department of Molecular Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205.
Kauffman
M G
MG
Kelly
T J
TJ
D000469:Alkaline Phosphatase / Q000378:metabolism; D000595:Amino Acid Sequence; D000957:Antigens, Viral, Tumor / Q000378:metabolism; D002384:Catalysis; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis*; D006367:HeLa Cells; D006801:Humans; D007399:Interphase; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000502:physiology*; D010766:Phosphorylation; D054648:Protein Phosphatase 2; D013539:Simian virus 40 / Q000235:genetics*
D000957:Antigens, Viral, Tumor; D004279:DNA, Viral; D000469:Alkaline Phosphatase; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2