Vaccinia virus G9 protein is an essential component of the poxvirus entry-fusion complex.
The vaccinia virus G9R gene (VACWR087) encodes a protein of 340 amino acids with the following structural features that are conserved in all poxviruses: a site for N-terminal myristoylation, 14 cysteines, and a C-terminal transmembrane domain. Previous studies showed that G9 is one of eight proteins associated in a putative entry-fusion complex. Our attempt to isolate a mutant without the G9R gene was unsuccessful, suggesting that it is essential for virus replication. To further investigate its role, we constructed a recombinant vaccinia virus in which G9R is regulated by addition of an inducer. Induced G9 protein was associated with mature infectious virions and could be labeled with a membrane-impermeant biotinylation reagent, indicating surface exposure. Omission of inducer reduced the infectious-virus yield by about 1.5 logs; nevertheless, all stages of virus morphogenesis appeared normal and extracellular virions were present on the cell surface. Purified virions assembled without inducer had a specific infectivity of less than 5% of the normal level and a comparably small amount of G9, whereas their overall polypeptide composition, including other components of the entry-fusion complex, was similar to that of virions made in the presence of inducer or of wild-type virions. G9-deficient virions bound to cells, but penetration of cores into the cytoplasm and early viral RNA synthesis were barely detected, and cell-cell fusion was not triggered by low pH. Of the identified components of the multiprotein complex, G9 is the sixth that has been shown to be required for entry and membrane fusion.
Ojeda
Suany
S
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892-0445, USA.
Domi
Arban
A
Moss
Bernard
B
D000595:Amino Acid Sequence; D002459:Cell Fusion; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D015967:Gene Expression Regulation, Viral / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D011485:Protein Binding; D016415:Sequence Alignment; D014158:Transcription, Genetic / Q000235:genetics; D014616:Vaccinia virus / Q000737:chemistry / Q000145:classification / Q000502:physiology* / Q000648:ultrastructure; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014779:Virus Replication
D014764:Viral Proteins; D007544:Isopropyl Thiogalactoside