Abstract
Subversion of the host immune system by viruses is often mediated by molecular decoys that sequester host proteins pivotal to mounting effective immune responses. The widespread mammalian pathogen parapox Orf virus deploys GIF, a member of the poxvirus immune evasion superfamily, to antagonize GM-CSF (granulocyte macrophage colony-stimulating factor) and IL-2 (interleukin-2), two pleiotropic cytokines of the mammalian immune system. However, structural and mechanistic insights into the unprecedented functional duality of GIF have remained elusive. Here we reveal that GIF employs a dimeric binding platform that sequesters two copies of its target cytokines with high affinity and slow dissociation kinetics to yield distinct complexes featuring mutually exclusive interaction footprints. We illustrate how GIF serves as a competitive decoy receptor by leveraging binding hotspots underlying the cognate receptor interactions of GM-CSF and IL-2, without sharing any structural similarity with the cytokine receptors. Our findings contribute to the tracing of novel molecular mimicry mechanisms employed by pathogenic viruses.
Highlights
Subversion of the host immune system by viruses is often mediated by molecular decoys that sequester host proteins pivotal to mounting effective immune responses
Via a series of cross-validated structural snapshots including the crystal structure of GM-CSF/IL-2 inhibition factor (GIF) in complex with granulocyte macrophage colony-stimulating factor (GM-CSF), two-dimensional (2D) classifications of GIF:GM-CSF and GIF:IL-2 complexes by negative-stain electron microscopy, the ensuing three-dimensional (3D) reconstruction of the GIF:IL-2 complex, and modeling of small-angle X-ray scattering (SAXS) data in conjunction with multi-angle laser light scattering (MALLS) measurements, we show that GIF adopts an obligate dimeric structure that can bind two molecules of its target cytokines
Addition of a molar excess of either recombinant ovine GM-CSF or IL-2 to culture medium containing secreted recombinant GIF enabled in situ reconstitution of GIF:GM-CSF and GIF:IL-2 complexes that could be purified to homogeneity for further biophysical and structural studies
Summary
Subversion of the host immune system by viruses is often mediated by molecular decoys that sequester host proteins pivotal to mounting effective immune responses. The Orf virus subverts the immune response by secreting virulence proteins that inhibit or mimic key anti-inflammatory host effector molecules[15], including a chemokine-binding protein (CKBP)[16], an Orf virus homologue of interleukin-10 (orfIL-10)[17] and vascular endothelial growth factor (VEGF-E)[18], an interferon resistance protein (OVIFNR) resembling the vaccinia virus E3L gene[19] and a secreted dual inhibitor of both granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin-2 (IL-2), termed GM-CSF/IL-2 inhibition factor (GIF)[20] The latter, GIF, is arguably one of the most intriguing viral proteins known because it targets two distinct host proteins pivotal to the host’s ability to mount an effective immune response. The structural and mechanistic basis for the remarkable functional duality displayed by GIF has remained uncharacterized
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