FIG. 1.

Construction and characterization of a recombinant NDV vector expressing MV H. (A) Schematic representation of the NDV-H cDNA construct. The hemagglutinin gene from the MV Schwarz vaccine strain (MV H) was inserted between the P and M genes of the molecular clone of rNDV/F3aa as described in Materials and Methods. (B) Expression of MV H protein in infected cells. Vero cells mock infected or infected with NDV-H were harvested 48 h postinfection, and the cell lysates were analyzed for MV H protein expression by Western blotting. (C) MV H expression by immunostaining of infected Vero cells. Vero cells were infected with NDV-H or measles virus; 24 h after infection, the cells were fixed and stained with anti-NDV polyclonal antibodies (red) and MV H monoclonal antibodies (green). DAPI staining revealed nuclear chromatin (blue). The arrow indicates a measles virus-infected syncytium. (D) Measles virus hemagglutinin is incorporated into the NDV virion. Wild-type NDV or recombinant viruses expressing RFP, MV H, or influenza A virus hemagglutinin were purified once or twice over a sucrose gradient. Virus proteins were separated by SDS-PAGE and stained with NDV- and MV-specific antisera (not shown) or Coomassie blue. The positions of the NDV hemagglutinin/neuraminidase (HN), nucleoprotein (NP), fusion protein (F1), polymerase (P), and matrix protein (M) are indicated on the right. Measles virus hemagglutinin is indicated by an asterisk. (E) MV H-specific antibodies block NDV-H expression. NDV-RFP and NDV-H were preincubated with neutralizing antibodies specific for MV H at different neutralizing titers and used to infect Vero cells. Virus-infected cells were visualized 48 h later by immunofluorescence.