Lyn and Fyn function as molecular switches that control immunoreceptors to direct homeostasis or inflammation.

Immunoreceptors can transduce either inhibitory or activatory signals depending on ligand avidity and phosphorylation status, which is modulated by the protein kinases Lyn and Fyn. Here we show that Lyn and Fyn control immune receptor signaling status. SHP-1 tyrosine 536 phosphorylation by Lyn activates the phosphatase promoting inhibitory signaling through the immunoreceptor. By contrast, Fyn-dependent phosphorylation of SHP-1 serine 591 inactivates the phosphatase, enabling activatory immunoreceptor signaling. These SHP-1 signatures are relevant in vivo, as Lyn deficiency exacerbates nephritis and arthritis in mice, whereas Fyn deficiency is protective. Similarly, Fyn-activating signature is detected in patients with lupus nephritis, underlining the importance of this Lyn-Fyn balance. These data show how receptors discriminate negative from positive signals that respectively result in homeostatic or inflammatory conditions.Src-family kinases Fyn and Lyn are signaling components downstream of ITAM-bearing antigen receptors. Here the authors show that by phosphorylating SHP-1 at different residues, Lyn and Fyn can have opposing regulatory effects on ITAM receptors.

Negative regulation of bacterial killing and inflammation by two novel CD16 ligands.

Sepsis, a leading cause of death worldwide, involves exacerbated proinflammatory responses and inefficient bacterial clearance. Phagocytic cells play a crucial part in the prevention of sepsis by clearing bacteria through host innate receptors. Here, we used a phage display library to identify two peptides in Escherichia coli that interact with host innate receptors. One of these peptides, encoded by the wzxE gene of E. coli K-12, was involved in the transbilayer movement of a trisaccharide-lipid intermediate in the assembly of enterobacterial common antigen. Peptide-receptor interactions induced CD16-mediated inhibitory immunoreceptor tyrosine-based activating motif signaling, blocking the production of ROS and bacterial killing. This CD16-mediated inhibitory signaling was abrogated in a WzxE(-/-) mutant of E. coli K-12, restoring the production of ROS and bacterial killing. Taken together, the two novel CD16 ligands identified negatively regulate bacterial killing and inflammation. Our findings may contribute toward the development of new immunotherapies for E. coli-mediated infectious diseases and inflammation.

IgA, IgA receptors, and their anti-inflammatory properties.

Immunoglobulin A (IgA) is the most abundantly produced antibody isotype in mammals. The primary function of IgA is to maintain homeostasis at mucosal surfaces and play a role in immune protection. IgA functions mainly through interaction with multiple receptors including IgA Fc receptor I (FcαRI), transferrin receptor 1 (CD71), asialoglycoprotein receptor (ASGPR), Fcα/µR, FcRL4, and DC-SIGN/SIGNR1. In this review we discuss recent data demonstrating anti-inflammatory functions of IgA through two receptors, the FcαRI and DC-SIGN/SIGNR1 interactions in the regulation of immunity. Serum monomeric IgA is able to mediate an inhibitory signal following the interaction with FcαRI. It results in partial phosphorylation of its FcRγ-ITAM and the recruitment of the tyrosine phosphatase SHP-1, which induces cell inhibition following the formation of intracellular clusters named inhibisomes. In contrast, cross-linking of FcαRI by multimeric ligands induces a full phosphorylation of the FcRγ-ITAM leading to the recruitment of the tyrosine kinase Syk and cell activation. In addition, secretory IgA can mediate a potent anti-inflammatory function following the sugar-dependent interaction with SIGNR1 on dendritic cells which induces an immune tolerance via regulatory T cell expansion. Overall, the anti-inflammatory effect of serum and secretory IgA plays a crucial role in the physiology and in the prevention of tissue damage in multiple autoimmune and inflammatory diseases.

Differential activation of Toll-like receptor-mediated apoptosis induced by hypoxia.

Ischemia-reperfusion injury induces intense inflammatory response and tissue damages resulting from the capacity of endogenous constituents called damageassociated molecular patterns (DAMPs) released by damaged or necrotic cells, to activate signaling pathways mediated by receptors of the innate immune systems. Among them, two members of the Toll-like receptors (TLR) family, TLR2 and TLR4 have been shown to play key roles in the induction of inflammatory response and cell apoptosis in a variety of ischemic tissues. The oxidative stress injury caused by I/R injury has been attributed to the activation of MAP kinase pathways, including those of ERK, JNK and p38. Here, we summarise recent findings concerning the role of the protein phosphatase 5 involved in the selective regulation of TLR2-mediated ERK1/2 signaling and the identification of the key role of the non-phagocytic NADPH oxidase 4 producing reactive oxygen species in the control of TLR4-mediated apoptosis in murine models of renal I/R injury and in post-hypoxic kidney tubule cells. The identification of molecules signaling involved in the ER stress-induced apoptotic signaling cascade may therefore represent potential targets to prevent the induction of apoptosis in hypoxic tissues.

Clostridium septicum alpha-toxin forms pores and induces rapid cell necrosis.

Alpha-toxin is the unique lethal virulent factor produced by Clostridium septicum, which causes traumatic or non-traumatic gas gangrene and necrotizing enterocolitis in humans. Here, we analyzed channel formation of the recombinant septicum alpha-toxin and characterized its activity on living cells. Recombinant septicum alpha-toxin induces the formation of ion-permeable channels with a single-channel conductance of about 175pS in 0.1M KCl in lipid bilayer membranes, which is typical for a large diffusion pore. Septicum alpha-toxin channels remained mostly in the open configuration, displayed no lipid specificity, and exhibited slight anion selectivity. Septicum alpha-toxin caused a rapid decrease in the transepithelial electrical resistance of MDCK cell monolayers grown on filters, and induced a rapid cell necrosis in a variety of cell lines, characterized by cell permeabilization to propidium iodide without DNA fragmentation and activation of caspase-3. Septicum alpha-toxin also induced a rapid K(+) efflux and ATP depletion. Incubation of the cells in K(+)-enriched medium delayed cell death caused by septicum alpha-toxin or epsilon-toxin, another potent pore-forming toxin, suggesting that the rapid loss of intracellular K(+) represents an early signal of pore-forming toxins-mediated cell necrosis.

Heat shock protein gp96 interacts with protein phosphatase 5 and controls toll-like receptor 2 (TLR2)-mediated activation of extracellular signal-regulated kinase (ERK) 1/2 in post-hypoxic kidney cells.

Ischemia/reperfusion injury (IRI) induces an innate immune response, leading to an inflammatory reaction and tissue damage that have been attributed to engagement of the Toll-like receptor (TLR) 2 and 4. However, the respective roles of TLR2 and/or TLR4 in mediating downstream activation of mitogen-activated protein kinase (MAPK) pathways during IRI have not been fully elucidated. Here we show that extracellular signal-regulated kinase (ERK)1/2 is activated in both intact kidneys and cultured renal tubule epithelial cells (RTECs) from wildtype and Tlr4 knockout mice, but not those from Tlr2 knockout mice subjected to transient ischemia. Geldanamycin (GA), an inhibitor of heat shock protein 90 and reticulum endoplasmic-resident gp96, and gp96 mRNA silencing (siRNA), did not affect ERK1/2 activation in either post-hypoxic wild-type or Tlr4-deficient RTECs, but did restore its activation in post-hypoxic Tlr2-deficient RTECs. Immunoprecipitation studies revealed that gp96 co-immunoprecipitates with the serine-threonine protein phosphatase 5 (PP5), identified as a negative modulator of the mitogen extracellular kinase (MEK)-ERK pathway, in unstressed wild-type and post-hypoxic Tlr2-deficient RTECs. In contrast, PP5 co-immunoprecipitation with gp96 was strikingly reduced in post-hypoxic wild-type RTECs, suggesting that the inactivation of PP5 resulting from the dissociation of PP5 from gp96 allows the activation of ERK1/2 to occur. Inhibition of PP5 by okadaic acid, and Pp5 siRNA also restored TLR2-mediated phosphorylation of ERK1/2, and apoptosis signal-regulating kinase 1 (ASK1)/c-Jun N-terminal kinase (JNK)-mediated apoptosis in post-hypoxic Tlr2-deficient RTECs. These findings indicate that gp96 interacts with PP5 and controls TLR2-mediated induction of ERK1/2 in post-hypoxic renal tubule cells.

NOX enzymes and Toll-like receptor signaling.

Invading microorganisms are recognized by the host innate immune system through pattern recognition receptors. Among these receptors, Toll-like receptors (TLRs) are able to sense the molecular signatures of microbial pathogens, protozoa, fungi, and virus and activate proinflammatory signaling cascades. In addition to their role in bacterial killing by phagocytes, reactive oxygen species generated by NADPH oxidase (NOX) homologues also play key roles in signaling and host defense in a variety of cell types. Recent studies have demonstrated a link between TLR activation and NOX homologues following microbial recognition highlighting their important role in the innate immune response and host defense.

Reduced expression of the NADPH oxidase NOX4 is a hallmark of adipocyte differentiation.

Adipocyte differentiation is a complex process regulated among other factors by insulin and the production of reactive oxygen species (ROS). NOX4 is a ROS generating NADPH oxidase enzyme mediating insulin's action in 3T3L1 adipocytes. In the present paper we show that NOX4 is expressed at high levels both in white and brown preadipocytes and that differentiation into adipocytes results in a decrease in their NOX4 mRNA content. These in vitro results were confirmed in vivo by demonstrating that in intact adipose tissue the majority of NOX4 expressing cells are localized within the preadipocyte containing stromal/vascular fraction, rather than in the portion consisting of mature adipocytes. In line with these observations, quantification of NOX4 mRNA in fat derived from different rodent models of insulin resistance indicated that alteration in NOX4 expression reflects changes in the ratio of adipocyte/interstitial fractions. In conclusion, we reveal that decreased NOX4 mRNA content is a hallmark of adipocyte differentiation and that NOX4 expression measured in whole adipose tissue is not an unequivocal indicator of intact or impaired insulin action.