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Mol Cell Biol. 1982 June; 2(6): 666-673

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

¹ Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario, Canada M5G 1L6

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 -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 -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 -amanitin can be transferred to TsAF8 cells and that the TS defect in TsAF8 is a polymerase II mutation.