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

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Abstract
Two conditional transcriptase-negative mutants of vesicular sotmatitis virus (VSV) serotype New Jersey, tsB1 and tsF1, their revertants tsB1/R1 and tsF1/R1 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.degree. C) and restrictive (39.degree. 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 30.degree. C only when the L fractions of tsB1 or tsF1 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 tsF1 and support previous observations that polypeptide L is the transciptase itself. The fact that a 2nd mutant of complementation group F, tsF2, is transcriptase-positive but replicase-negative suggests that polypeptide L is involved both in trascription and replication. Intracistronic complementations may account for the observation that the temperature-sensitive mutations affect polypeptide L in complementation groups B and F.