Smallpox vaccines induce antibodies to the immunomodulatory, secreted vaccinia virus complement control protein

Vaccination with Dryvax elicits a broad humoral response against many viral proteins. Human vaccinia immune globulin was used to screen the secreted proteins from cells infected with Dryvax or the candidate smallpox vaccine LC16m8 to determine whether the protective humoral response included antibodies against secreted viral proteins. Many proteins were detected, with the primary band corresponding to a band of 28 or 30 kDa in cells infected with Dryvax or LC16m8, respectively. This was identified as the vaccinia virus complement protein (VCP), which migrated more slowly in LC16m8-infected cells due to post-translational glycosylation. Vaccinia virus deleted in VCP, vVCPko, protected mice from a lethal intranasal challenge of vaccinia Western Reserve strain. Mice vaccinated with purified VCP demonstrated a strong humoral response, but were not protected against a moderate lethal challenge of vaccinia virus, suggesting that the humoral response against VCP is not critical for protection.

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The vaccinia virus A56 protein: a multifunctional transmembrane glycoprotein that anchors two secreted viral proteins

Journal of General Virology ◽

10.1099/vir.0.030460-0 ◽

2011 ◽

Vol 92 (9) ◽

pp. 1971-1980 ◽

Cited By ~ 7

Author(s):

Brian C. DeHaven ◽

Kushol Gupta ◽

Stuart N. Isaacs

Keyword(s):

Vaccinia Virus ◽

Protein A ◽

Viral Proteins ◽

Gene Encoding ◽

Biologically Relevant ◽

Complement Control Protein ◽

Transmembrane Glycoprotein ◽

Infected Cells ◽

Foreign Genes ◽

Control Protein

The vaccinia virus A56 protein was one of the earliest-described poxvirus proteins with an identifiable activity. While originally characterized as a haemagglutinin protein, A56 has other functions as well. The A56 protein is capable of binding two viral proteins, a serine protease inhibitor (K2) and the vaccinia virus complement control protein (VCP), and anchoring them to the surface of infected cells. This is important; while both proteins have biologically relevant functions at the cell surface, neither one can locate there on its own. The A56–K2 complex reduces the amount of virus superinfecting an infected cell and also prevents the formation of syncytia by infected cells; the A56–VCP complex can protect infected cells from complement attack. Deletion of the A56R gene results in varying effects on vaccinia virus virulence. In addition, since the gene encoding the A56 protein is non-essential, it can be used as an insertion point for foreign genes and has been deleted in some viruses that are in clinical development as oncolytic agents.

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Cell Surface Expression of the Vaccinia Virus Complement Control Protein Is Mediated by Interaction with the Viral A56 Protein and Protects Infected Cells from Complement Attack

Journal of Virology ◽

10.1128/jvi.02426-07 ◽

2008 ◽

Vol 82 (9) ◽

pp. 4205-4214 ◽

Cited By ~ 28

Author(s):

Natasha M. Girgis ◽

Brian C. DeHaven ◽

Xin Fan ◽

Kendra M. Viner ◽

Mohammad Shamim ◽

...

Keyword(s):

Membrane Protein ◽

Cell Surface ◽

Vaccinia Virus ◽

Transmembrane Domain ◽

Surface Expression ◽

Cell Surface Expression ◽

Major Protein ◽

Complement Control Protein ◽

Infected Cells ◽

Control Protein

ABSTRACT The vaccinia virus (VACV) complement control protein (VCP) is the major protein secreted from VACV-infected cells. It has been reported that VCP binds to the surfaces of uninfected cells by interacting with heparan sulfate proteoglycans (HSPGs). In this study, we show that VCP is also expressed on the surfaces of infected cells and demonstrate that surface localization occurs independently of HSPGs. Since VCP does not contain a transmembrane domain, we hypothesized that VCP interacts with a membrane protein that localizes to the infected-cell surface. We show that the VACV A56 membrane protein is necessary for the cell surface expression of VCP and demonstrate that VCP and A56 interact in VACV-infected cells. Since the surface expression of VCP was abrogated by reducing agents, we examined the contribution of an unpaired cysteine residue on VCP to VCP surface expression and VCP's interaction with A56. To do this, we mutated the unpaired cysteine in VCP and generated a recombinant virus expressing the altered form of VCP. Following the infection of cells with the mutant virus, VCP was neither expressed on the cell surface nor able to interact with A56. Importantly, the cell surface expression of VCP was found to protect infected cells from complement-mediated lysis. Our findings suggest a new function for VCP that may be important for poxvirus pathogenesis and impact immune responses to VACV-based vaccines.

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