A Temporal Proteomic Map of Epstein-Barr Virus Lytic Replication in B Cells

Epstein-Barr virus (EBV) replication contributes to multiple human diseases, including infectious mononucleosis, nasopharyngeal carcinoma, B cell lymphomas, and oral hairy leukoplakia. We performed systematic quantitative analyses of temporal changes in host and EBV proteins during lytic replication to gain insights into virus-host interactions, using conditional Burkitt lymphoma models of type I and II EBV infection. We quantified profiles of >8,000 cellular and 69 EBV proteins, including >500 plasma membrane proteins, providing temporal views of the lytic B cell proteome and EBV virome. Our approach revealed EBV-induced remodeling of cell cycle, innate and adaptive immune pathways, including upregulation of the complement cascade and proteasomal degradation of the B cell receptor complex, conserved between EBV types I and II. Cross-comparison with proteomic analyses of human cytomegalovirus infection and of a Kaposi-sarcoma-associated herpesvirus immunoevasin identified host factors targeted by multiple herpesviruses. Our results provide an important resource for studies of EBV replication

Signal Peptide-Dependent Inhibition of MHC Class I Heavy Chain Translation by Rhesus Cytomegalovirus

The US2-11 region of human and rhesus cytomegalovirus encodes a conserved family of glycoproteins that inhibit MHC-I assembly with viral peptides, thus preventing cytotoxic T cell recognition. Since HCMV lacking US2-11 is no longer able to block assembly and transport of MHC-I, we examined whether this is also observed for RhCMV lacking the corresponding region. Unexpectedly, recombinant RhCMV lacking US2-11 was still able to inhibit MHC-I expression in infected fibroblasts, suggesting the presence of an additional MHC-I evasion mechanism. Progressive deletion analysis of RhCMV-specific genomic regions revealed that MHC-I expression is fully restored upon additional deletion of rh178. The protein encoded by this RhCMV-specific open reading frame is anchored in the endoplasmic reticulum membrane. In the presence of rh178, RhCMV prevented MHC-I heavy chain (HC) expression, but did not inhibit mRNA transcription or association of HC mRNA with translating ribosomes. Proteasome inhibitors stabilized a HC degradation intermediate in the absence of rh178, but not in its presence, suggesting that rh178 prevents completion of HC translation. This interference was signal sequence-dependent since replacing the signal peptide with that of CD4 or murine HC rendered human HCs resistant to rh178. We have identified an inhibitor of antigen presentation encoded by rhesus cytomegalovirus unique in both its lack of homology to any other known protein and in its mechanism of action. By preventing signal sequence-dependent HC translocation, rh178 acts prior to US2, US3 and US11 which attack MHC-I proteins after protein synthesis is completed. Rh178 is the first viral protein known to interfere at this step of the MHC-I pathway, thus taking advantage of the conserved nature of HC leader peptides, and represents a new mechanism of translational interference

The human cytomegalovirus ul11 protein interacts with the receptor tyrosine phosphatase cd45, resulting in functional paralysis of t cells

Human cytomegalovirus (CMV) exerts diverse and complex effects on the immune system, not all of which have been attributed to viral genes. Acute CMV infection results in transient restrictions in T cell proliferative ability, which can impair the control of the virus and increase the risk of secondary infections in patients with weakened or immature immune systems. In a search for new immunomodulatory proteins, we investigated the UL11 protein, a member of the CMV RL11 family. This protein family is defined by the RL11 domain, which has homology to immunoglobulin domains and adenoviral immunomodulatory proteins. We show that pUL11 is expressed on the cell surface and induces intercellular interactions with leukocytes. This was demonstrated to be due to the interaction of pUL11 with the receptor tyrosine phosphatase CD45, identified by mass spectrometry analysis of pUL11-associated proteins. CD45 expression is sufficient to mediate the interaction with pUL11 and is required for pUL11 binding to T cells, indicating that pUL11 is a specific CD45 ligand. CD45 has a pivotal function regulating T cell signaling thresholds; in its absence, the Src family kinase Lck is inactive and signaling through the T cell receptor (TCR) is therefore shut off. In the presence of pUL11, several CD45-mediated functions were inhibited. The induction of tyrosine phosphorylation of multiple signaling proteins upon TCR stimulation was reduced and T cell proliferation was impaired. We therefore conclude that pUL11 has immunosuppressive properties, and that disruption of T cell function via inhibition of CD45 is a previously unknown immunomodulatory strategy of CMV

A Vibrio parahaemolyticus T3SS Effector Mediates Pathogenesis by Independently Enabling Intestinal Colonization and Inhibiting TAK1 Activation.

Vibrio parahaemolyticus type III secretion system 2 (T3SS2) is essential for the organism's virulence, but the effectors required for intestinal colonization and induction of diarrhea by this pathogen have not been identified. Here, we identify a type III secretion system (T3SS2)-secreted effector, VopZ, that is essential for V. parahaemolyticus pathogenicity. VopZ plays distinct, genetically separable roles in enabling intestinal colonization and diarrheagenesis. Truncation of VopZ prevents V. parahaemolyticus colonization, whereas deletion of VopZ amino acids 38-62 abrogates V. parahaemolyticus-induced diarrhea and intestinal pathology but does not impair colonization. VopZ inhibits activation of the kinase TAK1 and thereby prevents the activation of MAPK and NF-κB signaling pathways, which lie downstream. In contrast, the VopZ internal deletion mutant cannot counter the activation of pathways regulated by TAK1. Collectively, our findings suggest that VopZ's inhibition of TAK1 is critical for V. parahaemolyticus to induce diarrhea and intestinal pathology

A Vibrio parahaemolyticus T3SS Effector Mediates Pathogenesis by Independently Enabling Intestinal Colonization and Inhibiting TAK1 Activation.

Vibrio parahaemolyticus type III secretion system 2 (T3SS2) is essential for the organism's virulence, but the effectors required for intestinal colonization and induction of diarrhea by this pathogen have not been identified. Here, we identify a type III secretion system (T3SS2)-secreted effector, VopZ, that is essential for V. parahaemolyticus pathogenicity. VopZ plays distinct, genetically separable roles in enabling intestinal colonization and diarrheagenesis. Truncation of VopZ prevents V. parahaemolyticus colonization, whereas deletion of VopZ amino acids 38-62 abrogates V. parahaemolyticus-induced diarrhea and intestinal pathology but does not impair colonization. VopZ inhibits activation of the kinase TAK1 and thereby prevents the activation of MAPK and NF-κB signaling pathways, which lie downstream. In contrast, the VopZ internal deletion mutant cannot counter the activation of pathways regulated by TAK1. Collectively, our findings suggest that VopZ's inhibition of TAK1 is critical for V. parahaemolyticus to induce diarrhea and intestinal pathology

Cytokine receptor IL27RA is an NF-kB-responsive gene involved in CD38 upregulation in multiple myeloma

Multiple myeloma (MM) shows constitutive activation of canonical and non-canonical nuclear factor-ĸB (NF-ĸB) signaling through genetic mutations or stimuli from the tumour microenvironment (TME). A subset of MM cell lines showed dependency for cell growth and survival on the canonical NF-ĸB transcription factor RELA alone, suggesting a critical role for a RELA-mediated biological program in MM pathogenesis. Here, we determined the RELA-dependent transcriptional program in MM cell lines and found the expression of the cell surface molecules IL-27 receptor-α (IL-27Rα) and the adhesion molecule JAM2 to be responsive to RELA at the mRNA and protein levels. IL-27Rα and JAM2 were expressed on primary MM cells at higher levels than on normal long-lived plasma cells (PCs) in the bone marrow. IL-27 activated STAT1, and to a lesser extent STAT3, in MM cell lines and in PCs generated from memory B-cells in an IL-21-dependent in vitro PC-differentiation assay. Concomitant activity of IL-21 and IL-27 enhanced differentiation into PCs and increased cell-surface expression of the known STAT target gene CD38. In accordance, a subset of MM cell lines and primary MM cells cultured with IL-27 upregulated CD38 cell-surface expression, a finding with potential implications for enhancing the efficacy of CD38-directed monoclonal antibody (mAb) therapies by increasing CD38-expression on tumour cells. The elevated expression of IL-27Rα and JAM2 on MM cells compared to normal PCs may be exploited for the development of targeted therapeutic strategies that modulate the interaction of MM cells with the TME

Structural mechanism of ER retrieval of MHC class I by cowpox.

One of the hallmarks of viral immune evasion is the capacity to disrupt major histocompatibility complex class I (MHCI) antigen presentation to evade T-cell detection. Cowpox virus encoded protein CPXV203 blocks MHCI surface expression by exploiting the KDEL-receptor recycling pathway, and here we show that CPXV203 directly binds a wide array of fully assembled MHCI proteins, both classical and non-classical. Further, the stability of CPXV203/MHCI complexes is highly pH dependent, with dramatically increased affinities at the lower pH of the Golgi relative to the endoplasmic reticulum (ER). Crystallographic studies reveal that CPXV203 adopts a beta-sandwich fold similar to poxvirus chemokine binding proteins, and binds the same highly conserved MHCI determinants located under the peptide-binding platform that tapasin, CD8, and natural killer (NK)-receptors engage. Mutagenesis of the CPXV203/MHCI interface identified the importance of two CPXV203 His residues that confer low pH stabilization of the complex and are critical to ER retrieval of MHCI. These studies clarify mechanistically how CPXV203 coordinates with other cowpox proteins to thwart antigen presentation