Involvement of the influenza A virus PB2 protein in the regulation of viral gene expression.

To determine the function(s) of the PB2 protein of influenza A virus, six temperature-sensitive (ts) mutants of A/Udorn/72 (H3N2) virus, each carrying a ts mutation in the PB2 gene, were analysed for virus RNA and protein synthesis. One of the mutants, ICRC27, exhibited unique phenotypes and was characterized in detail. At the non-permissive temperature, 40 degrees C, the accumulation of mRNA for each genome segment was reduced severely, leading to delayed and reduced synthesis of viral proteins, complementary and viral RNAs (cRNAs and vRNAs). At the permissive temperature, 34 degrees C, the mutant virus produced several-fold greater concentrations of both mRNAs and cRNAs of PB2, PB1 and PA segments than wild-type virus. The synthesis of the three polymerase proteins and the induction of RNA polymerase activity were also greatly increased. By contrast, the expression of the haemagglutinin (HA) gene was severely suppressed. The over-production of the polymerase mRNAs was not observed during primary transcription, i.e. in the presence of cycloheximide. The ts+ revertants of ICRC27 did not exhibit the ts defects and also lost most of the non-ts phenotypes at 34 degrees C. These observations indicate that the PB2 protein participates not only in the synthesis of viral RNAs, but also in the regulation of viral gene expression, i.e. in the down-regulation of the three polymerase genes and the up-regulation of the HA gene during secondary transcription.