Expression of the influenza virus haemagglutinin in insect cells by a baculovirus vector.
The insect baculovirus Autographa californica nuclear polyhedrosis virus (AcNPV) has played a major role in studies on the molecular biology of insect DNA viruses. Recently, this system has been effectively adapted as a highly efficient vector in insect cells for the expression of several mammalian genes. A cDNA sequence of the influenza (fowl plague) virus haemagglutinin gene has been inserted into the BamHI site of the pAc373 polyhedrin vector. Spodoptera frugiperda cells were co-transfected with this construct, pAc-HA651, and authentic AcNPV DNA. Recombinant virus was selected by adsorption of transfected cells to erythrocytes followed by serial plaque passages on S. frugiperda cells. We have determined the site of insertion of the haemagglutinin gene into the AcNPV genome by restriction enzyme cleavage and Southern blot hybridization analyses using haemagglutinin cDNA as a probe. The influenza haemagglutinin gene is located in the polyhedrin gene of AcNPV DNA. Immunofluorescent labelling, immunoprecipitation and immunoblot analyses with specific antisera revealed that S. frugiperda cells produce immune reactive haemagglutinin after infection with the recombinant virus. The haemagglutinin is expressed at the cell surface and has haemolytic capacity that has been activated by post-translational proteolytic cleavage. When chickens were immunized with S. frugiperda cells expressing haemagglutinin, they developed haemagglutinin-inhibiting and neutralizing antibodies and were protected from infection with fowl plague virus. These observations demonstrate that the haemagglutinin is processed in insect cells in a similar fashion as in fowl plaque virus-infected vertebrate cells and that it has full biological activity.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004262:DNA Restriction Enzymes; D005455:Fluorescent Antibody Technique; D005796:Genes; D005814:Genes, Viral; D005822:Genetic Vectors; D006386:Hemagglutination Tests; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral / Q000235:genetics*; D006461:Hemolysis; D007304:Insect Viruses / Q000235:genetics*; D007700:Kinetics; D010957:Plasmids
D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral; D004262:DNA Restriction Enzymes