DNA-Mediated Transfer of an RNA Polymerase II Gene: Reversion of the Temperature-Sensitive Hamster Cell Cycle Mutant TsAF8 by Mammalian DNA

Abstract

Treatment of the TsAF8 temperature-sensitive (TS) mutant of Syrian hamster BHK-21 cells, with calcium phosphate precipitates of genomic TS + DNAs from a variety of mammalian cell lines permitted the selection of TS + colonies at 40°C. TS + transformation events were distinguished from spontaneous TS + reversions in experiments in which α-amanitin-sensitive (Ama S ) TS + DNA was used to transform an Ama R derivative of TsAF8 cells and Ama R TS + DNA was used to transform Ama S TsAF8 cells. In each case it was possible to demonstrate the unselected acquisition of the appropriate Ama S or Ama R phenotype with the selected TS + allele. Each of these TS + transformed cell lines when grown at 40°C contained an RNA polymerase II activity with a sensitivity to inhibition by α-amanitin characteristic of the particular DNA used to transform the TS cells, whereas at 34°C the same cells contained a mixture of Ama R and Ama S polymerase II activities. Together, these data provide convincing evidence that the RNA polymerase II gene determining sensitivity to inhibition by α-amanitin can be transferred to TsAF8 cells and that the TS defect in TsAF8 is a polymerase II mutation.