Identification of endoprotease activity in the trans Golgi membranes of rat liver cells that specifically processes in vitro the fusion glycoprotein precursor of virulent Newcastle disease virus.
A ubiquitous host endoprotease(s) responsible for activation of the fusion glycoprotein precursor (F0) of virulent Newcastle disease virus (NDV) is an important determinant for its spreading and organ tropism in the host. To characterize the virus-activating protease (VAP), we isolated endoprotease activity from the trans Golgi membranes of rat liver cells by using F0-containing NDV particles grown in a lymphoid cell line NALM6 as substrate. The enzyme cleaved in vitro only the F0 protein of virulent NDV but not that of an avirulent strain, suggesting that it specifically recognizes pairs of basic residues at the cleavage site. Furthermore, the enzyme was found to be membrane-bound, calcium ion-dependent, and active over a broad pH range, from 6 to 8. The inhibitor spectrum of the protease together with the enzyme properties described above indicates that it is a KEX2-like enzyme. Experiments using monensin, A23187, and chloroquine indicate that the F0 cleavage of virulent NDV occurs normally in rat primary hepatocytes at or before the trans Golgi and is a calcium-dependent process. The correspondence between the characteristics of the cleavage in rat hepatocytes and those of the rat protease in vitro indicates that the endoprotease is a strong candidate for the VAP that determines the pantropic nature of virulent NDV.
Sakaguchi
T
T
Department of Bacteriology, Hiroshima University School of Medicine, Japan.
Matsuda
Y
Y
Kiyokage
R
R
Kawahara
N
N
Kiyotani
K
K
Katunuma
N
N
Nagai
Y
Y
Yoshida
T
T
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology; D002413:Cations, Divalent; D002460:Cell Line; D010450:Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D006056:Golgi Apparatus / Q000201:enzymology / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008970:Molecular Weight; D009522:Newcastle disease virus / Q000378:metabolism* / Q000472:pathogenicity; D011092:Polyethylene Glycols / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D051381:Rats; D014760:Viral Fusion Proteins / Q000378:metabolism*
D002413:Cations, Divalent; D011480:Protease Inhibitors; D014760:Viral Fusion Proteins; D000001:Calcimycin; D011092:Polyethylene Glycols; D010450:Endopeptidases; D002118:Calcium