oligomycins and Cell-Transformation--Neoplastic

A common metabolic change in cancer is the acquisition of glycolytic phenotypes. Increased expression of glycolytic enzymes is considered as one contributing factor. The role of mitochondrial defects in acquisition of glycolytic phenotypes has been postulated but remains controversial. Here we show that functional defects in mitochondrial respiration could be induced by oncogenic H-Ras(Q61L) transformation, even though the mitochondrial contents or mass was not reduced in the transformed cells. First, mitochondrial respiration, as measured by mitochondrial oxygen consumption, was suppressed in NIH-3T3 cells transformed with H-Ras(Q61L). Second, oligomycin or rotenone did not reduce the cellular ATP levels in the H-Ras(Q61L) transformed cells, suggesting a diminished role of mitochondrial respiration in the cellular energy metabolism. Third, inhibition of glycolysis with iodoacetic acid reduced ATP levels at a much faster rate in H-Ras(Q61L) transformed cells than in the vector control cells. The reduction of cellular ATP levels was reversed by exogenously added pyruvate in the vector control cells but not in H-Ras(Q61L) transformed cells. Finally when compared to the HRas(Q61L) transformed cells, the vector control cells had increased resistance toward glucose deprivation. The increased resistance was dependent on mitochondrial oxidative phosphorylation since rotenone or oligomycin abolished the increased survival of the vector control cells under glucose deprivation. The results also suggest an inability of the H-Ras(Q61L) transformed cells to reactivate mitochondrial respiration under glucose deprivation. Taken together, the data suggest that mitochondrial respiration can be impaired during transformation of NIH-3T3 cells by oncogeneic H-Ras(Q61L).

Topics: Adenosine Triphosphate; Animals; Antimycin A; Cell Transformation, Neoplastic; Electron Transport; Energy Metabolism; Fibroblasts; Genes, ras; Glucose; Glycolysis; Iodoacetic Acid; Mice; Mitochondria; Mutation, Missense; NIH 3T3 Cells; Oligomycins; Oncogene Protein p21(ras); Oxidative Phosphorylation; Oxygen Consumption; Point Mutation; Pyruvic Acid; Rotenone

Topics: Animals; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cell Line; Cell Transformation, Neoplastic; Embryo, Mammalian; Fibroblasts; Methylcholanthrene; Mice; Mice, Inbred C3H; Oligomycins; Oxygen Consumption; Phorbols; Tetradecanoylphorbol Acetate

Normal rat kidney cells infected with a temperature-sensitive mutant (LA23) of Rous sarcoma virus exhibit the transformed phenotype when grown at 33 degrees and the normal phenotype at 39 degrees. We have previously shown [Ash, J.F., Vogt, P.K. & Singer, S.J. (1976) Proc. Natl. Acad. Sci. USA 73, 3603-3607] that the addition of protein synthesis inhibitors to LA23-infected cells grown at 33 degrees causes them to revert, over a period of 12 hr, to the normal phenotype with respect to morphological and cytoskeletal characteristics. We now show that reversion of the metabolic characteristics of the transformed phenotype to those of the normal also occurs under these conditions. LA23-infected cells show an increased rate of aerobic glycolysis at 33 degrees compared to that at 39 degrees. They also show a different sensitivity of that rate to dinitrophenol and oligomycin at 33 degrees compared to 39 degrees. Such cells grown at 33 degrees in the presence of cycloheximide or abrin rapidly recover the aerobic glycolysis characteristics of the normal phenotype. These results support the thesis that transformation by the src gene of the Rous sarcoma virus is a pleiotypic and reversible process, such as is involved in a pleiotypic enzymic modification reaction and its reversal.

Topics: Aerobiosis; Avian Sarcoma Viruses; Cell Transformation, Neoplastic; Cell Transformation, Viral; Cells, Cultured; Dinitrophenols; Glycolysis; Oligomycins; Temperature; Time Factors

Topics: Adenosine Triphosphate; Agglutination; Azides; Cell Aggregation; Cell Line; Cell Transformation, Neoplastic; Concanavalin A; Dinitrophenols; Glucose; Glycine max; Iodoacetates; Kinetics; Lectins; Oligomycins; Plant Lectins; Receptors, Drug; Temperature

Transformed fibroblasts had a low content of ATP when grown at a high cell density and a high content of ATP when grown at a low cell density. Concanavalin A agglutinated transformed cells with a low, but not those with a high, ATP content. Transformed cells with a high ATP content gained agglutinability after ATP depletion by inhibitors of the energy-generating systems, and those with a low ATP content lost their agglutinability after restoration of a high ATP content by glucose. Fixation of the surface membrane by formaldehyde, glutaraldehyde, or LaCl(3), inhibited agglutination of cells with an ATP content that allows agglutination. Normal fibroblasts grown at a high or a low cell density were not agglutinated by concanavalin A. Depletion of the cellular ATP content of normal cells induced agglutination only in cells grown at a high, but not at a low, cell density. A similar number of concanavalin A molecules was bound to the surface membrane of agglutinating and nonagglutinating fibroblasts. It is suggested that a high content of ATP inhibits the movement of concanavalin A binding sites, and that a low content of ATP allows, in transformed cells, a new distribution of binding sites to form the clusters required for cell agglutination. Agglutinability of transformed cells is determined by ATP content, and in normal cells changes in the content of ATP are by themselves not sufficient to induce agglutination. Transformed cells, therefore, do not have a control, presumably for membrane stability, that exists in normal cells.

Topics: Adenosine Triphosphate; Agglutination; Aldehydes; Animals; Binding Sites; Cell Line; Cell Membrane; Cell Transformation, Neoplastic; Concanavalin A; Cricetinae; Dinitrophenols; Fibroblasts; Fluorides; Formaldehyde; Glucose; Hydrazones; Lanthanum; Nitrites; Oligomycins; Polyomavirus; Simian virus 40