Vaccinia virus A17L open reading frame encodes an essential component of nascent viral membranes that is required to initiate morphogenesis.
We generated an antiserum to the predicted C-terminal peptide of the A17L open reading frame (ORF), which encodes a 23-kDa polypeptide with hydrophobic regions characteristic of membrane proteins. Immuno-electron microscopy of infected cells indicated that the A17L protein is intimately associated with the earliest characteristic viral membranes, even those formed in the presence of the drug rifampin. To study the role of the A17L protein in morphogenesis, we constructed recombinant vaccinia viruses in which the endogenous A17L ORF was deleted and a copy of the ORF under the control of the bacteriophage T7 RNA polymerase and the Escherichia coli lac repressor was inserted into an alternative site in the vaccinia virus genome. Growth of these recombinant viruses was entirely dependent on the induction of A17L expression by isopropyl-beta-D-thiogalactopyranoside. Electron microscopic examination of cells infected in the absence of inducer revealed the accumulation of large, well-demarcated electron-dense aggregates but no characteristic membrane-associated viral structures. Viral late protein synthesis occurred under these conditions, although the maturational proteolytic processing of structural proteins was inhibited. We conclude that the product of the A17L gene is an essential component of the immature viral membrane and has an early function in viral morphogenesis.
Wolffe
E J
EJ
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, Bethesda, Maryland 20892, USA.
Moore
D M
DM
Peters
P J
PJ
Moss
B
B
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002460:Cell Line; D002522:Chlorocebus aethiops; D017931:DNA Primers; D004279:DNA, Viral / Q000378:metabolism; D005822:Genetic Vectors; D006367:HeLa Cells; D014645:Herpesvirus 3, Human / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D008565:Membrane Proteins; D008854:Microscopy, Electron; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D009024:Morphogenesis / Q000187:drug effects; D016366:Open Reading Frames; D010957:Plasmids; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D014759:Viral Envelope Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D010948:Viral Plaque Assay; D014779:Virus Replication
C081257:A17L protein, Vaccinia virus; D017931:DNA Primers; D004279:DNA, Viral; D008565:Membrane Proteins; D014759:Viral Envelope Proteins; D007544:Isopropyl Thiogalactoside