Orf Virus Encodes a Novel Secreted Protein Inhibitor of Granulocyte-Macrophage Colony-Stimulating Factor and Interleukin-2

Abstract

The parapoxvirus orf virus encodes a novel soluble protein inhibitor of ovine granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-2 (IL-2). The GM-CSF- and IL-2-inhibitory factor (GIF) gene was expressed as an intermediate-late viral gene in orf virus-infected cells. GIF formed homodimers and tetramers in solution, and it bound ovine GM-CSF with a Kd of 369 pM and ovine IL-2 with a Kd of 1.04 nM. GIF did not bind human GM-CSF or IL-2 in spite of the fact that orf virus is a human pathogen. GIF was detected in afferent lymph plasma draining the skin site of orf virus reinfection and was associated with reduced levels of lymph GM-CSF. GIF expression by orf virus indicates that GM-CSF and IL-2 are important in host antiviral immunity.

Conservation and variation of the parapoxvirus GM-CSF-inhibitory factor (GIF) proteins

“…On: Fri, 11 May 2018 21:58:45 GIF (rGIF) proteins were selected by assaying for GIF activity using a modification of the ovine (ov) GM-CSF-specific ELISA (Entrican et al, 1996;Deane et al, 2000). The pEE14 vector contains a glutamine synthetase (GS) gene; therefore, cell lines transfected stably with the plasmid can be selected on the basis of their survival in methionine sulphoximine (MSX; Sigma).…”

“…The presence of biologically active ORFV GIF can be measured indirectly by its ability to interfere with the detection of ovine/bovine GM-CSF by specific ELISA. Cell-free supernates and cell lysates from GIF cDNA-transfected and virus-infected cells were assayed for GM-CSF-binding activity as described previously (Deane et al, 2000). Binding of GIF to ovine/bovine IL-2 was similarly measured by ELISA.…”

“…The integrity of the genes was verified by sequencing prior to transfection into COS-7 fibroblasts or CHO cells. Stable cell lines expressing each of the GIF proteins were established as described previously (Deane et al, 2000). Cell lines expressing the recombinant ORFV and PCPV IP: 34.214.212.150…”

“…All three viruses can infect humans. As is common amongst other poxviruses, ORFV has been shown to encode factors that either mimic host immunoregulatory proteins (Fleming et al, 1997;Haig & Fleming, 1999;Seet et al, 2003) or have the potential for interacting with components of the host immune system (Haig et al, 1998;McInnes et al, 1998McInnes et al, , 2005Deane et al, 2000). In doing so, it is thought that these virus-encoded factors are capable of subverting the host immune response to infection, creating an environment suitable for efficient virus replication.…”

“…Some, such as the vFLIP protein (MC159) of molluscum contagiosum virus, which inhibits apoptosis, may be species-specific (Shisler & Moss, 2001;Thurau et al, 2006). ORFV has been shown to encode a protein (GIF) that is capable of binding and inhibiting both of the ovine cytokines interleukin-2 (IL-2) and granulocytemacrophage colony-stimulating factor (GM-CSF) (Deane et al, 2000;McInnes et al, 2005). Such a protein has not been found so far in any other poxvirus; we therefore investigated whether it was conserved in other PPVs.…”