The replication activity of influenza virus polymerase is linked to the capacity of the PA subunit to induce proteolysis

The PA subunit of the influenza virus polymerase complex is a phosphorylate d protein that induces a proteolytic process that decreases its own accumul ation levels and those of coexpressed proteins, The aminoterminal third of the protein is responsible for the induction of proteolysis. We mutated fiv e potential casein kinase II phosphorylation sites located in the amino-ter minal third of the protein. Mutations affecting position 157 almost complet ely abrogated proteolysis induction, whereas a mutation at position 162 pro duced a moderate decrease and mutations at positions 151, 200, and 223 did not affect proteolysis induction. Reconstitution of the influenza virus pol ymerase in vivo with viral model RNA containing the chloramphenicol acetylt ransferase (CAT) gene indicated that the CAT activity obtained correlated w ith the capacity of each PA mutant to induce proteolysis, RNA protection as says of the products obtained with viral polymerase, reconstituted in vivo with model RNAs, indicated that mutations at position 157 led to a selectiv e loss of the ability to synthesize cRNA from the viral RNA template but no t to transcribe viral RNA while a mutation affecting position 162 shelved a n intermediate phenotype, Collectively, these data provide a link between P A-mediated induction of proteolysis and the replication activity of the pol ymerase.