Relationship between DNA topoisomerase I activity and DNA synthesis in cultured hepatocytes: effects of orotic acid.

P. Giannoni, G. Fronza, L. Scarabelli, E. Fugassa, M. Orunesu, C. F. Cesarone

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Previous studies have demonstrated that DNA topoisomerase I activity can be closely related to DNA replication and active transcription in different experimental models. This relationship was further investigated by studying the time course of DNA topoisomerase I activity in cultured rat hepatocytes stimulated by epidermal growth factor. This mitogen has been shown to stimulate DNA synthesis in liver cells both in vivo and in vitro. DNA topoisomerase I activity was assayed quantifying ATP-independent relaxation of a negatively supercoiled plasmid, substrate for the enzyme. Incubation of cellular extracts was carried out at 30 degrees C for 15'. Products were electrophoresed and analyzed by densitometry. DNA synthesis was measured as [3H]thymidine incorporation in the hepatocytes, after a 2 hrs pulse labelling. An increase in DNA topoisomerase I activity was observed early after epidermal growth factor addition, before the onset of DNA synthesis. Concomitant administration of orotic acid, which has recently shown mitoinhibitory effect, abolished the EGF-induced activity as well as DNA synthesis. Taken together these data indicate that, in cultured hepatocytes, the induction of DNA synthesis is supported by DNA topoisomerase I with a precise time schedule. Orotic acid administration abolishes this response and causes an overall loss in DNA topoisomerase I activity.

Original languageEnglish
Pages (from-to)509-516
Number of pages8
JournalBollettino della Societa Italiana di Biologia Sperimentale
Issue number5
Publication statusPublished - May 1991

ASJC Scopus subject areas

  • Medicine(all)


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