novobiocin and Cell-Transformation--Neoplastic

A differentiation inducer butyrate and a tumor promoter teleocidin had inhibitory effects on the proliferation of PLC/PRF/5 hepatoma. Both of these reagents stimulated the production of procollagen type III peptide, enhanced the cytokeratin assembly and altered the morphological appearance. Novobiocin, a topoisomerase II inhibitor, enhanced the cytokeratin assembly induced by butyrate but antagonized that induced by teleocidin without changing the expression and the phosphorylation state of cytokeratin proteins. In addition, novobiocin acted synergistically with butyrate but not with teleocidin in stimulating the procollagen production and the acetate uptake. These results suggest that butyrate and teleocidin induce cell differentiation via distinct signaling pathway and that novobiocin and butyrate can be used as subsidiary drugs in preventing the growth of hepatoma.

Topics: Acetylation; Butyrates; Butyric Acid; Carcinogens; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Drug Synergism; Humans; Keratins; Liver Neoplasms; Lyngbya Toxins; Novobiocin; Peptide Fragments; Procollagen; Tumor Cells, Cultured

The activity of topoisomerase II and the cellular content of the 170kD and 180kD forms of the enzyme were studied as functions of transformation and growth state by using normal and ras-transformed NIH-3T3 cells. Total topoisomerase II activity, as measured by the unknotting of P4 DNA, was higher in ras-transformed than in normal cells in similar growth states, and was higher in exponentially growing than in plateau cells for both cell lines. Total topoisomerase II levels, as measured by immunoblotting, showed a similar dependence on transformation and growth state. The relative amounts of the 170kD and 180kD forms of the enzyme varied as a function of transformation and growth state. The proportion of 170kD topoisomerase II was higher in ras-transformed than in untransformed cells and depended much less on growth state in the ras-transformed cells. The topoisomerase II activity in extracts of ras-transformed cells was more sensitive to inhibition by teniposide and merbarone, drugs which selectively inhibit the 170kD form of topoisomerase II. The ras-transformed cells were also more sensitive to the cytotoxic effects of these drugs. An increase in the relative cellular content of 170kD topoisomerase II is characteristic of ras-transformed 3T3 cells, and the levels of this form of the enzyme appear to be less dependent on proliferation state than in untransformed cells. The susceptibility of certain tumors to killing by topoisomerase II-directed drugs may be due to a higher proportion of 170kD enzyme as well as a higher level of total topoisomerase II activity.

Topics: Animals; Blotting, Northern; Blotting, Western; Cell Division; Cell Line; Cell Nucleus; Cell Transformation, Neoplastic; DNA Damage; DNA Topoisomerases, Type II; Genes, ras; Mice; Molecular Weight; Novobiocin; RNA, Messenger; Teniposide; Thiobarbiturates; Topoisomerase II Inhibitors

The activities of topoisomerases I and II were assayed in subcellular extracts obtained from nontumorigenic BALB/c 3T3 A31 and normal rat kidney (NRK) cell lines and from the same cells transformed by benzo[a]pyrene (BP-A31), Moloney (M-MSV-A31) and Kirsten (K-A31) sarcoma viruses, and simian virus 40 (SV-NRK). The enzymatic activity of topoisomerase I was monitored by the relaxation of negatively supercoiled pBR322 DNA and by the formation of covalent complexes between 32P-labeled DNA and topoisomerase I. Topoisomerase II activity was determined by decatenation of kinetoplast DNA (k-DNA). It was found that nuclear and cytoplasmic type I topoisomerase specific activities were higher in every transformed cell line than in the normal counterparts. These differences cannot be attributed to an inhibitory factor present in A31 cells. When chromatin was treated at increasing ionic strengths, the 0.4 M NaCl extract showed the highest topoisomerase I specific activity. Moreover, in this fraction the transformed cells exhibited the most significant increment in the enzymatic activity as compared with nontransformed cultures. Spontaneously transformed A31 cells showed topoisomerase I activity similar to that of extracts of cells transformed by benzo[a]pyrene. Topoisomerase II specific activity was also increased in SV-NRK cells, as judged by the assay for decatenation of k-DNA to yield minicircle DNA.

Topics: Animals; Benzo(a)pyrene; Cell Adhesion; Cell Nucleus; Cell Transformation, Neoplastic; Cell Transformation, Viral; DNA Topoisomerases, Type I; Mice; Novobiocin; Rats