Interaction of L and NS proteins of vesicular stomatitis virus with its template ribonucleoprotein during RNA synthesis in vitro.

Soluble transcriptase containing the L and the NS proteins was isolated from purified vesicular stomatitis virus and its binding with the template ribonucleoprotein containing the N protein-RNA complex was studied with respect to its ability to initiate and synthesize RNA in vitro. By using u.v.-irradiated template reconstituted with soluble transcriptase, it was shown that the synthesis of leader RNA and other small initiated mRNA sequences continued while full-length mRNA synthesis decreased by 90%. In the presence of ATP and CTP, the reconstituted complex synthesized polyphosphorylated oligonucleotides which include AC, AAC and AACA which represent 5'-terminal sequences transcribed from the leader template and genes coding for mRNAs. In the presence of arabinosyl ATP, an inhibitor of RNA synthesis in vitro, the synthesis of leader RNA was found to be inhibited considerably more than other small initiated mRNA sequences. Reconstitution of RNA synthesis with soluble transcriptase and template in the presence of viral matrix (M) protein at low ionic condition resulted in virtual cessation of leader RNA synthesis, although the synthesis of small initiated N mRNA, 11 to 14 bases, continued. These results suggest that transcriptase can bind at multiple sites on the genome template and initiate RNA chains.