Polypeptide composition of cell membranes from chick embryo fibroblasts transformed by rous sarcoma virus.
Chick embryo fibroblasts were transformed by the Bryan high-titer strain of Rous sarcoma virus (RSV-BH), or a mutant (RSV-BH-Ta) inducing temperature-dependent transformation. Surface membranes from normal and transformed cells were isolated as membrane vesicles by differential centrifugation, and as cell ghosts after ZnCl2 treatment and separation in an aqueous two-phase system. These preparations were analyzed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate or phenol/urea/acetic acid. In general a greater resolution of individual bands was found in gels containing phenol/urea/acetic acid, which separates polypeptides on the bases of size and charge. Electrophoresis of preparations from nontransformed cells showed that two polypeptides (molecular weights 200 000 and 250 000) found in cell ghosts were missing in membrane vesicles. In cell ghosts, transformation by RSV-BH resulted in a significant decrease of the 250 000 molecular weight complex. Also a polypeptide (molecular weight 73 000) prominent in membrane vesicles from nontransformed cells was decreased in transformed cells. Surfaces from cells transformed by RSV-BH-Ta at 37 degrees C presented patterns similar to those for RSV-BH infected cells. Shifting these cells to 41 degrees C resulted in an increase in the 250 000 molecular weight complex, although the amount of this protein(s) never reached that found in noninfected cells. Inhibitors of RNA and protein synthesis failed to block the morphological changes occurring in RSV-BH-Ta cells after temperature shifts from 41 degrees C to 37 degrees C or vice-versa. The same inhibitors caused a reduction in the levels of the 250 000 molecular weight complex at both temperatures. These data indicate that these large membrane-associated polypeptides play little or no role in the morphological changes associated with transformation and its reversal.
D000818:Animals; D001358:Avian Sarcoma Viruses; D002462:Cell Membrane / Q000378:metabolism; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002642:Chick Embryo; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D009154:Mutation; D011506:Proteins / Q000378:metabolism*; D013696:Temperature
D011506:Proteins; D003609:Dactinomycin; D003513:Cycloheximide