A little cooperation helps murine cytomegalovirus (MCMV) go a long way: MCMV co-infection rescues a chemokine salivary gland defect.

Cytomegaloviruses (CMVs) produce chemokines (vCXCLs) that have both sequence and functional homology to host chemokines. Assessment of vCXCL-1's role in CMV infection is limited to in vitro and in silico analysis due to CMVs species specificity. In this study, we used the murine CMV (MCMV) mouse model to evaluate the function of vCXCL-1 in vivo. Recombinant MCMVs expressing chimpanzee CMV vCXCL-1 (vCXCL-1CCMV) or host chemokine, mCXCL1, underwent primary dissemination to the popliteal lymph node, spleen and lung similar to the parental MCMV. However, neither of the recombinants expressing chemokines was recovered from the salivary gland (SG) at any time post-infection although viral DNA was detected. This implies that the virus does not grow in the SG or the overexpressed chemokine induces an immune response that leads to suppressed growth. Pointing to immune suppression of virus replication, recombinant viruses were isolated from the SG following infection of immune-ablated mice [i.e. SCID (severe combined immunodeficiency), NSG (non-obese diabetic SCID gamma) or cyclophosphamide treated]. Depletion of neutrophils or NK cells does not rescue the recovery of chemokine-expressing recombinants in the SG. Surprisingly we found that co-infection of parental virus and chemokine-expressing virus leads to the recovery of the recombinants in the SG. We suggest that parental virus reduces the levels of chemokine expression leading to a decrease in inflammatory monocytes and subsequent SG growth. Therefore, aberrant expression of the chemokines induces cells of the innate and adaptive immune system that curtail the growth and dissemination of the recombinants in the SG.

Modulation of innate immune signaling by the secreted form of the West Nile virus NS1 glycoprotein.

West Nile virus (WNV) employs several different strategies to escape the innate immune response. We have previously demonstrated that the WNV NS1 protein interferes with signal transduction from Toll-like receptor 3 (TLR3). NS1 is a glycoprotein that can be found intracellularly or associated with the plasma membrane. In addition, NS1 is secreted to high levels during flavivirus infections. We investigated whether the secreted form of NS1 inhibits innate immune signaling pathways in uninfected cells. Secreted NS1 (sNS1) was purified from supernatants of cells engineered to express the protein. Purified sNS1 associated with and repressed TLR3-induced cytokine production by HeLa cells, and inhibited signaling from TLR3 and other TLRs in bone marrow-derived macrophages and dendritic cells. Footpad administration of sNS1 showed the protein associated predominantly with macrophages and dendritic cells in the draining lymph node. Additionally, sNS1 significantly reduced TLR3 signaling and WNV replicon particle-mediated cytokine transcription in popliteal lymph nodes.

Intrinsic adjuvanting of a novel single-cycle flavivirus vaccine in the absence of type I interferon receptor signaling.

Type I interferons (IFNs) are critical for controlling pathogenic virus infections and can enhance immune responses. Hence their impact on the effectiveness of live-attenuated vaccines involves a balance between limiting viral antigen expression and enhancing the development of adaptive immune responses. We examined the influence of type I IFNs on these parameters following immunization with RepliVAX WN, a single-cycle flavivirus vaccine (SCFV) against West Nile virus (WNV) disease. RepliVAX WN-immunized mice produced IFN-α and displayed increased IFN-stimulated gene transcription in draining lymph nodes (LN). SCFV gene expression was over 100 fold-higher on days 1-3 post-infection in type I IFN receptor knockout mice (IFNAR(-/-)) compared to wild-type (wt) mice indicating a profound IFN-mediated suppression of SCFV gene expression in the wt animals. IFNAR(-/-) mice produced nearly equivalent levels of WNV-specific serum IgG and WNV-specific CD4(+) T cell responses compared to wt mice. However, significantly higher numbers of WNV-specific CD8(+) T cells were produced by IFNAR(-/-) mice and a significantly greater percentage of these T cells from IFNAR(-/-) mice produced only IFN-γ following antigen-specific re-stimulation. This altered cytokine expression was not associated with increased antigen load suggesting the loss of type I IFN receptor signaling was responsible for the altered quality of the CD8(+) effector T cell response. Together, these results indicate that although type I IFN is not essential for the intrinsic adjuvanting of RepliVAX WN, it plays a role in shaping the cytokine secretion profiles of CD8(+) effector T cells elicited by this SCFV.

Feeling manipulated: cytomegalovirus immune manipulation.

No one likes to feel like they have been manipulated, but in the case of cytomegalovirus (CMV) immune manipulation, we do not really have much choice. Whether you call it CMV immune modulation, manipulation, or evasion, the bottom line is that CMV alters the immune response in such a way to allow the establishment of latency with lifelong shedding. With millions of years of coevolution within their hosts, CMVs, like other herpesviruses, encode numerous proteins that can broadly influence the magnitude and quality of both innate and adaptive immune responses. These viral proteins include both homologues of host proteins, such as MHC class I or chemokine homologues, and proteins with little similarity to any other known proteins, such as the chemokine binding protein. Although a strong immune response is launched against CMV, these virally encoded proteins can interfere with the host's ability to efficiently recognize and clear virus, while others induce or alter specific immune responses to benefit viral replication or spread within the host. Modulation of host immunity allows survival of both the virus and the host. One way of describing it would be a kind of "mutually assured survival" (as opposed to MAD, Mutually Assured Destruction). Evaluation of this relationship provides important insights into the life cycle of CMV as well as a greater understanding of the complexity of the immune response to pathogens in general.

Functional characterization of chimpanzee cytomegalovirus chemokine, vCXCL-1(CCMV).

Human cytomegaloviruses (HCMVs) are important pathogens in immunocompromised patients and newborns. The viral chemokine, vCXCL-1, of the Toledo (Tol) strain of HCMV has been implicated in HCMV virulence. Chimpanzee CMV (CCMV) has several genes with similarity to the vCXCL-1(Tol) gene, UL146. In order to test whether the CCMV viral chemokine, vCXCL-1(CCMV), is similar to vCXCL-1(Tol), we characterized its function in vitro. Receptor binding, activation, chemotaxis, signaling, and apoptosis in neutrophils were compared between vCXCL-1(Tol) and vCXCL-1(CCMV) and host chemokines. Although the homologues had similar activation potentials, chemotactic properties, and signaling, vCXCL-1(CCMV) had a approximately 70-fold lower affinity for CXCR2 and displayed differences in integrin upregulation and neutrophil apoptosis. These data demonstrate that in spite of extensive amino acid variability in vCXCL-1, CCMV may provide a model for assessing the role of vCXCL-1 in CMV pathogenesis in vivo.