An inhibitor of the in vitro elongation reaction of simian virus 40 DNA replication is overcome by proliferating-cell nuclear antigen

S. H. Lee, Y. Ishimi, M. K. Kenny, P. Bullock, F. B. Dean, J. Hurwitz

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Abstract

The replication of simian virus 40 (SV40) origin-containing DNA has been reconstituted by using SV40 large tumor (T) antigen and cellular proteins purified from HeLa cells. This replication is unaffected by proliferating-cell nuclear antigen (PCNA). In contrast, PCNA has been reported to stimulate SV40 DNA synthesis carried out with crude fractions [Prelich, G., Kostura, M., Marshak, D.R., Mathews, M.B. & Stillman, B. (1987) Nature (London) 326, 471-475]. This difference is caused by the presence of a protein in crude fractions that inhibits the elongation of nascent DNA chains during replication. In the presence of PCNA, crude fractions containing this elongation inhibition factor can extend DNA chains. We describe the partial purification of this inhibitor and show that its addition limited SV40 DNA replication to the synthesis of short chains, an effect reversed by the addition of PCNA. However, the reversal of the inhibition by PCNA in the SV40 system required additional protein fractions distinct from PCNA and the enzymes constituting the purified system. These results suggest that the PCNA-mediated effect on SV40 DNA replication may be indirect. Such an interplay between negative and positive regulatory functions including PCNA may contribute to the control of DNA synthesis characteristic of the eukaryotic cell cycle.

Original languageEnglish (US)
Pages (from-to)9469-9473
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume85
Issue number24
DOIs
StatePublished - Jan 1 1988

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