Several residues within the N-terminal arm of vesicular stomatitis virus nucleoprotein play a critical role in protecting viral RNA from nuclease digestion

The nucleoprotein (N) of vesicular stomatitis virus (VSV) plays a central role in transcription and replication by encapsidating genome RNA to form a nucleocapsid as the template for the RNA synthesis. Using minigenome system we evaluated the roles of 21 amino acids of the N-terminal arm of N in forming functional N-RNA templates and found that three triple-amino-acid substitutions (TVK4-6A3, RII7-9A3, and VIV13-15A3) and one single-amino-acid substitution (R7A) resulted in RNA synthesis loss. But all the mutants maintain the ability to oligomerize N, interact with P, and encapsidate viral RNA for template formation. Further analysis showed that the nucleocapsid formed by these mutants failed to protect RNA from nuclease digestion. Then, we found that only recombinant viruses containing R7A could be recovered. Our results show that the several amino acids within the N-terminal arm of N contribute to the template function beyond its role in RNA encapsidation and viral growth.