STUDIES OF THE STRUCTURAL PROTEINS OF NEWCASTLE DISEASE VIRUS

Part I. A unique abundant protein (P) analogous to the putative polymerase proteins of other paramyxoviruses was identified in Newcastle disease virus (NDV). Monomers and disulfide-linked trimers of the protein were separated from other viral proteins using nonreducing sodium dodecyl sulfate-polyacrylamide gels. Four structural forms of P having different isoelectric points were resolved by two-dimensional electrophoresis. A fifth species was found in infected cells and was synthesized in vitro. The P proteins were found associated with nucleocapsids using fractionation, radioiodination and immunochemical techniques. Two virion forms of P were phosphorylated as were the nucleocapsid and nonglycosylated membrane proteins.^ Dimers of the hemagglutinin-neuraminidase (HN) glycoprotein, two disulfide-linked versions of the fusion glycoprotein and several electrophoretic variants of the nucleocapsid protein were isolated from virions under nonreducing conditions.^ Radioisotopic pulse-chase experiments and polyacrylamide gel analyses were used to study the kinetics of assembly of polypeptides into virions. Newly synthesized core proteins, including each of the four forms of P, were rapidly assembled into virions whereas the appearance of envelope glycoproteins was delayed. Proteins synthesized during a short labeling period were available for assembly into virions for several hours except for P polypeptides which were rapidly depleted.^ Part II. A temperature-sensitive (ts) mutant (C1) of NDV with defective envelope glycoproteins was characterized by revertant analysis. Low hemagglutination and neuraminidase activities of C1 were due to metabolically unstable HN polypeptides which were assembled into virions in reduced amounts. C1 was also ts for cytopathology, had low virulence for chicken embryos and had nonhemolytic and exceptionally thermolabile virions.^ A revertant that formed small plaques at the nonpermissive temperature recovered hemolytic activity, thermostability and normal cytopathology for cells but retained the HN and low virulence defects. A unique second-step large plaque revertent (L1) was isolated that assembled wild-type (wt) amounts of stable HN polypeptides in virions and exhibited wt levels of hemagglutination but had less than 3% of the neuraminidase activity. The lack of neuraminidase did not impair reproduction of L1 in cultured cells nor its virulence in ovo but did lower efficiencies of attachment and elution. The attachment defect was due to the presence of sialic acid on virions.