EXPRESSION AND CHARACTERIZATION OF THE INFLUENZA-A VIRUS NONSTRUCTURAL PROTEIN NS1 IN YEAST

The influenza A virus non-structural protein NS1 was produced using a copper-inducible expression system in the yeast Saccharomyces cerevisi ae. The protein produced had a molecular weight of 26 kDa by SDS-PAGE and was reactive with anti-NS1 antisera. The recombinant NS1 protein w as targetted to the nucleolus/nuclear envelope fraction of the yeast c ell nucleus, showing that its localisation signals remain functional i n yeast. In addition, immune-electron microscopy detected cytoplasmic inclusions reminiscent of those seen in cells infected with some influ enza strains. The NS1 protein was shown to be capable of in vivo self- interaction which probably forms the basis of its propensity to form i nclusions. Expression of the protein was found to be toxic to yeast ce lls expressing it, supporting a role for the protein in the shutdown o f influenza virus-infected cells. Deletion mapping of NS1 pointed to 2 regions of the molecule being important for this toxicity: a basic C- terminal stretch which has been shown to act as a nuclear localisation signal, and an N-terminal region implicated in RNA binding.