Temperature Sensitivity of the Transcriptase of Mutants tsB1 and tsF1 of Vesicular Stomatitis Virus New Jersey Is a Consequence of Mutation

Two condition~il transcriptase-negative mutants of vesicular stomatitis virus (VSV) serotype New Jersey, tsB 1 and tsF 1, their revertants B 1/R 1 and tsF 1/R 1 and the wild-type virus were dissociated into pellet, NS and L fractions and, after reconstitution of these in various combinations, the transcriptase activities were assayed in vitro at the permissive (31 °C) and restrictive (39 °C) temperatures. The pellet fractions contained the virion RNA-polypeptide N complexes, while the NS and L fractions were essentially pure preparations of these polypeptides. The synthesis of RNA by the reconstituted pellet and L fractions was inhibited at 39 °C only when the L fractions of tsB1 or tsFl were used. Addition of the NS fractions to the reconstituted pellet and L fractions did not alter the rates of RNA synthesis. These results demonstrate that polypeptide L is the temperature-sensitive polypeptide of both mutants tsB1 and tsFl and support previous observations that polypeptide L is the transcriptase itself. The fact that a second mutant of complementation group F, tsF2, is transcriptase-positive but replicase-negative suggests that polypeptide L is involved both in transcription and replication. Intracistronic complementations may account for the observation that the temperature-sensitive mutations affect polypeptide L in complementation groups B and F