TEMPERATURE-SENSITIVE PHENOTYPE OF THE HUMAN PARAINFLUENZA VIRUS TYPE-3 CANDIDATE VACCINE STRAIN (CP45) CORRELATES WITH A DEFECT IN THE L-GENE

We have previously demonstrated that the temperature sensitivity of a human parainfluenza virus type 3 (HPIV-3) candidate vaccine strain (cp 45), which is currently under evaluation in humans, is associated with poor transcriptional activity of the virus at the nonpermissive tempe rature (R. Ray, K. Meyer, F. Newman, and R. B. Belshe, J. Virol, 69:19 59-1963, 1995), In this study, the temperature sensitivity of cp45 vir us was further investigated by the complementation of a specific gene function, CV-1 cells were transfected with cloned genes from wild-type HPIV-3 encoding the large protein (L), phosphoprotein (P), and nucleo capsid protein (NP), alone or together, for the expression of biologic ally active proteins, Only cells expressing the L gene were able to re scue cp45 replication when incubated at the nonpermissive temperature (39.5 degrees C), whereas cells transiently expressing NP or P were in capable of rescuing the virus, The virus titers obtained following com plementation of the L protein were 190 to 2,300 PFU/ml of culture medi um, compared with the undetectable growth of the cp45 temperature-sens itive mutant at the nonpermissive temperature, Rescued progeny virus s till maintained the temperature-sensitive phenotype, Results from this study suggest that the temperature sensitivity of the cp45 candidate vaccine strain is associated primarily with L-protein function and tha t the defect can be complemented by transient expression of the wild-t ype protein, This study underscores the importance of the L protein in RNA polymerase activity and its critical role in virus replication.