STUDIES ON RABIES VIRUS-RNA POLYMERASE - 1 - CDNA CLONING OF THE CATALYTIC SUBUNIT (L PROTEIN) OF AVIRULENT HEP-FLURY STRAIN AND ITS EXPRESSION IN ANIMAL-CELLS

To investigate the RNA polymerase of rabies virus, we cloned a cDNA of the catalytic subunit (called L protein because of its large molecula r size) of the HEP-Flury strain, an avirulent strain obtained by high frequencies of serial embryonated hen egg passages. Nucleotide sequenc ing showed that the cDNA encodes a long polypeptide of 2,127 amino aci ds (Mr, 242,938), A comparison of the deduced amino acid sequence with that of other strains (PV and SAD B19) indicated that the sequence wa s highly conserved, except for several amino acid substitutions which were accumulated in some limited regions, A fragment of the cDNA was u sed for expression in Escherichia coli (E, coli) to prepare the L anti gen for raising the antibodies in rabbits. Immunoprecipitation studies with the rabbit antiserum showed that the polypeptides produced in th e L cDNA-transfected COS-7 cells displayed almost the same electrophor etic mobility as that of authentic L protein. Immunofluorescence studi es indicated that both L and P (another subunit of RNA polymerase) pro teins displayed colocalized distribution with the nucleocapsid antigen (N) in the cytoplasmic inclusion bodies, where envelope proteins (G a nd M) were absent. On the other hand, expression of the L protein alon e did not cause inclusion body-like granular distribution, suggesting that the inclusion body-like accumulation depends on certain interacti on(s) with other viral gene products, probably with the ribonucleoprot eins comprising the inclusion bodies.