RNA-POLYMERASE OF VESICULAR STOMATITIS-VIRUS SPECIFICALLY ASSOCIATES WITH TRANSLATION ELONGATION FACTOR-I ALPHA-BETA-GAMMA FOR ITS ACTIVITY

An RNA-dependent RNA polymerase is packaged within the virions of puri fied vesicular stomatitis virus, a nonsegmented negative-strand RNA vi rus, which carries out transcription of the genome RNA into mRNAs both in vitro and in vivo. The RNA polymerase is composed of two virally e ncoded polypeptides: a large protein L (240 kDa) and a phosphoprotein P (29 kDa). Recently, we obtained biologically active L protein from i nsect cells following infection by a recombinant baculovirus expressin g L gene. During purification of the L protein from Sf21 cells, we obt ained in addition to an active L fraction an inactive fraction that re quired uninfected insect cell extract to restore its activity. The cel lular factors have now been purified, characterized, and shown to be b eta and gamma subunits of the protein synthesis elongation factor EF-1 . We also demonstrate that the alpha subunit of EF-1 remains tightly b ound to the L protein in the inactive fraction and beta gamma subunits associate with the L(alpha) complex. Further purification of L(alpha) from the inactive fraction revealed that the complex is partially act ive and is significantly stimulated by the addition of beta gamma subu nits purified from Sf21 cells. A putative inhibitor(s) appears to co-e lute in the inactive fraction that blocked the L(alpha) activity. The purified virions also package all three subunits of EF-1. These findin gs have a striking similarity with Q beta RNA phage, which also associ ates with the bacterial homologue of EF-1 for its replicase function, implicating a possible evolutionary relationship between these host pr oteins and the RNA-dependent RNA polymerase of RNA viruses.