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Journal of Virology, March 2004, p. 2357-2366, Vol. 78, No. 5
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.5.2357-2366.2004

Vaccinia Virus Entry into Cells Is Dependent on a Virion Surface Protein Encoded by the A28L Gene

Tatiana G. Senkevich, Brian M. Ward,{dagger} and Bernard Moss*

Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892

Received 4 September 2003/ Accepted 27 October 2003

The A28L gene of vaccinia virus is conserved in all poxviruses and encodes a protein that is anchored to the surface of infectious intracellular mature virions (IMV) and consequently lies beneath the additional envelope of extracellular virions. A conditional lethal recombinant vaccinia virus, vA28-HAi, with an inducible A28L gene, undergoes a single round of replication in the absence of inducer, producing IMV, as well as extracellular virions with actin tails, but fails to infect neighboring cells. We show here that purified A28-deficient IMV appeared to be indistinguishable from wild-type IMV and were competent to synthesize RNA in vitro. Nevertheless, A28-deficient virions did not induce cytopathic effects, express early genes, or initiate a productive infection. Although A28-deficient IMV bound to the surface of cells, their cores did not penetrate into the cytoplasm. An associated defect in membrane fusion was demonstrated by the failure of low pH to trigger syncytium formation when cells were infected with vA28-HAi in the absence of inducer (fusion from within) or when cells were incubated with a high multiplicity of A28-deficient virions (fusion from without). The correlation between the entry block and the inability of A28-deficient virions to mediate fusion provided compelling evidence for a relationship between these events. Because repression of A28 inhibited cell-to-cell spread, which is mediated by extracellular virions, all forms of vaccinia virus regardless of their outer coat must use a common A28-dependent mechanism of cell penetration. Furthermore, since A28 is conserved, all poxviruses are likely to penetrate cells in a similar way.

* Corresponding author. Mailing address: Laboratory of Viral Diseases, National Institutes of Health, 4 Center Dr., MSC 0445, Bethesda, MD 20892-0445. Phone: (301) 496-9869. Fax: (301) 480-1147. E-mail: bmoss{at}nih.gov.

{dagger} Present address: Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, N.Y.

Journal of Virology, March 2004, p. 2357-2366, Vol. 78, No. 5
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.5.2357-2366.2004




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