Human CD317 is an intrinsic immunity factor that restricts the release of enveloped viruses, including the major pathogens HIV and Lassa virus, from infected cells in culture. Its importance for infection control in humans is unclear, due in part to its incompletely defined in vivo expression pattern. CD317 also has been proposed as a selective target for immunotherapy of multiple myeloma. To provide a framework for studies of the biological functions, regulation, and therapeutic potential of CD317, we performed microarray-based expression profiling in 468 tissue samples from 25 healthy organs from more than 210 patients. We found that CD317 protein was expressed to varying degrees in all organs tested and detected in a number of specialized cell types, including hepatocytes, pneumocytes, ducts of major salivary glands, pancreas and kidney, Paneth cells, epithelia, Leydig cells, plasma cells, bone marrow stromal cells, monocytes, and vascular endothelium. Although many of these cell types are in vivo targets for pathogenic viruses, restriction by CD317 or virus-encoded antagonists has been documented in only some of them. Limited cell type–dependent coexpression of CD317 with the IFN biomarker MxA in vivo and lack of responsive stimulation in organ explants suggest that interferons may only partially regulate CD317. This in vivo expression profiling sheds light on the biology and species-specificity of CD317, identifies multiple thus far unknown interaction sites of viruses with this restriction factor, and refutes the concept of its restricted constitutive expression and primary IFN inducibility. CD317's widespread expression calls into question its suitability as a target for immunotherapy.

Staphylococcus aureus contains three members of the LytR-CpsA-Psr (LCP) family of membrane proteins: MsrR, SA0908 and SA2103. The characterization of single-, double- and triple-deletion mutants revealed distinct phenotypes for each of the three proteins. MsrR was involved in cell separation and septum formation and influenced β-lactam resistance; SA0908 protected cells from autolysis; and SA2103, although displaying no apparent phenotype by itself, enhanced the properties of msrR and sa0908 mutants when deleted. The deletion of sa0908 and sa2103 also further attenuated the virulence of msrR mutants in a nematode-killing assay. The severely defective growth phenotype of the triple mutant revealed that LytR-CpsA-Psr proteins are essential for optimal cell division in S. aureus. Growth could be rescued to varying degrees by any one of the three proteins, indicating some functional redundancy within members of this protein family. However, differing phenotypic characteristics of all single and double mutants and complemented triple mutants indicated that each protein played a distinct role(s) and contributed differently to phenotypes influencing cell separation, autolysis, cell surface properties and virulence.

Induction of polyclonal B cell activation is a phenomenon observed in many types of infection, but its immunological relevance is unclear. In this study we show that staphylococcal protein A induces T cell–independent human B cell proliferation by enabling uptake of TLR-stimulating nucleic acids via the VH3+ BCR. We further demonstrate that Staphylococcus aureus strains with high surface protein A expression concomitantly trigger activation of human plasmacytoid dendritic cells (pDC). Sensitivity to chloroquine, cathepsin B inhibition, and a G-rich inhibitory oligodeoxynucleotide supports the involvement of TLR9 in this context. We then identify pDC as essential cellular mediators of B cell proliferation and Ig production in response to surface protein A–bearing S. aureus. The in vivo relevancy of these findings is confirmed in a human PBMC Nod/scidPrkdc/γc−/− mouse model. Finally, we demonstrate that co-operation of pDC and B cells enhances B cell–derived IL-10 production, a cytokine associated with immunosuppression and induction of IgG4, an isotype frequently dominating the IgG response to S. aureus. IL-10 release is partially dependent on TLR2-active lipoproteins, a hallmark of the Staphylococcus species. Collectively, our data suggest that S. aureus exploits pDC and TLR to establish B cell–mediated immune tolerance.

Suppressory B-cell function controls immune responses and is mainly dependent on IL-10 secretion. Pharmacological manipulation of B-cell-specific IL-10 synthesis could, thus, be therapeutically useful in B-cell chronic lymphocytic leukemia, transplantation, autoimmunity and sepsis. TLR are thought to play a protagonistic role in the formation of IL-10-secreting B cells. The aim of the study was to identify the molecular events selectively driving IL-10 production in TLR9-stimulated human B cells. Our data highlight the selectivity of calcineurin inhibitors in blocking TLR9-induced B-cell-derived IL-10 transcription and secretion, while IL-6 transcription and release, B-cell proliferation, and differentiation remain unaffected. Nevertheless, TLR9-induced IL-10 production was found to be independent of calcineurin phosphatase activity and was even negatively regulated by NFAT. In contrast to TLR9-induced IL-6, IL-10 secretion was highly sensitive to targeting of spleen tyrosine kinase ( Eur. J. Immunol. 2014. 44: 1285-1298 [7]. However, the mechanisms underlying this suppressory B-cell function were less well understood. To date, it is well accepted that the hallmark of B-cell regulatory function is the anti-inflammatory cytokine . IL-10-producing B cells arise in different contexts, among them B-cell chronic lymphocytic leukemia (B-CLL), infections, and the immune response to apoptotic cells [9][10][11]. They can limit ongoing T-cell responses [12]. While exclusive lack of TLR adaptor MyD88 in B lymphocytes leads to development of chronic EAE or aggravation of Salmonella infection [10], loss of B-cell-specific IL-10 drives autoimmunity in TLR9-deficient mice [13].Further reports propose that formation of plasma blasts coincides with release of 14]. This highlights the possibility that in B cells IL-10 production is triggered in an Ag-specific manner and might selectively target Ag-specific T cells. This would discriminate the function of B-cell-derived IL-10 from that produced by myeloid cells. As a consequence, specific pharmacological intervention with B-cell-derived IL-10 could represent a promising strategy to protect from destructive T-cell responses to both auto-and alloantigen while preserving immunity to infection.Transcriptional regulation of IL-10 synthesis is cell type specific [15]. In B cells, IL-10 production has been investigated upon BCR activation and in leukemia. The studies suggest a role of calcium sensors and NFAT transcription factors (TF) [11,16]. However, little is known on the regulation of B-cell-derived IL-10 production and in particular in response to TLR9 stimulation. The objective of the present study was to analyze the molecular mechanisms regulating IL-10 production in B cells challenged with TLR9 agonists. Results CpG oligodeoxynucleotides (ODN) and BCR cross-linking represent potent IL-10 inducersIn this study, we were interested in the molecular mechanisms mediating B-cell-derived release of IL-10 upon stimulation of TLR9. BCR-, TLR-, and CD40-signaling have all been implicate...

IRAK4, a serine/threonine kinase is a central adaptor protein in TLR signaling. To better understand the clinical significance of IRAK4 deficiency we examined the impact of IRAK4 on bacterial recognition in human monocytes. We show that IRAK4 knockdown modulates monocyte-derived cytokine secretion in response to Staphylococcus aureus and Streptococcus pneumoniae, resulting in decreased IL-12 and elevated IL-10 production, a finding also reproducible with ligands for TLR2 and TLR4. In contrast, silencing of MyD88 leads to a complete loss of cytokine secretion, indicating that IRAK4 acts as a differential regulator of bacteria/TLR-induced cytokine secretion downstream of MyD88. Further analysis revealed that this modulatory function results from IRAK4-mediated suppression of protein kinase B (PKB/Akt). Release of suppression upon IRAK4 silencing (but not MyD88 knockdown) increases phosphorylation of PKB/Akt, counteracts NF-κB activation and finally results in a monocyte phenotype with tolerogenic features, thus unleashing Akt-and mTOR-dependent release of IL-10, along with concomitant phosphorylation of FOXO transcription factors. In line with these observations IRAK4-deficient monocytes failed to induce allogeneic CD8 + and CD4 + T-cell responses, an effect reverted by neutralization of IL-10. Taken together, our data highlight an unexpected role of IRAK4, Akt, and mTOR in the regulation of tolerance in human monocytes.Keywords: IRAK4 r monocytes r PKB/Akt r TLR Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionMonocytes are among the first to encounter bacterial pathogens in infections of the bloodstream. They account for 10% of the human peripheral blood leukocytes, making monocytes one of the most Correspondence: Dr. Isabelle Bekeredjian-Ding e-mail: bekeredjian-ding@uni-bonn.de abundant antigen-presenting cell subsets in the circulation and a very potent source for cytokines [1,2]. Their ability to produce high levels of cytokines and thereby shape the systemic immune response is thought to be important in determining the outcome of sepsis and balancing pro-inflammatory versus compensatory anti-inflammatory responses [3].Human blood monocytes are a heterogeneous cell population and functionally defined subsets can be distinguished based on their cytokine and receptor expression profiles. Classical monocytes express high levels of CD14, but no CD16 (FcγRIII) C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu Eur. J. Immunol. 2013. 43: 1630-1642 Innate immunity 1631 and account for ∼90% of blood monocytes. The nonclassical subset is characterized by the expression of CD16 and low CD14 levels. While the classical CD14 + subset is characterized by the preferential production of anti-inflammatory IL-10 rather than pro-inflammatory cytokines after TLR stimulation, the nonclassical subset produces high amounts of TNF and low levels of IL-10 in response to TLR ligands, and is, therefore, referred to as proinflammatory [4][5][...

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