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Systemic lupus erythematosus etiology includes both genetic and environmental factors. Evidence suggests that many genetic loci in humans and mouse models contribute to the occurrence and clinical presentation of lupus. This large array of different genes affects many aspects of immune cell function, including the activation and functional differentiation of B cells, T cells, dendritic cells and other immune cells. In particular, the T-cell components that contribute to systemic lupus erythematosus pathogenesis are incompletely defined.A major paradigm shift in understanding how CD4+ T cells contribute to autoimmunity recently occurred with the discovery of a new T-cell population that produces the cytokine IL-17 (IL-17A), termed 'Th17'. Although Th17 cells contribute to autoimmune disease in rheumatoid arthritis and Crohn's disease, their role in systemic lupus erythematosus is far less clear.In this review, we focus on an emerging role for the cytokine IL-17 and the cells that produce it in contributing to lupus in particular based on recent findings in animal models.

The γc-family cytokine IL-2 activates signaling events that contribute to cell survival and proliferation, the best-studied of which are the STAT-5 and phosphatidylinositol 3-kinase (PI3K) pathways. The starting point of this study was to define genes regulated by the IL-2R-mediated PI3K pathway in T cells. Accordingly, we used an erythropoietin (EPO) receptor chimeric receptor system in which IL-2-dependent HT-2 T cells expressed a mutant EPO-IL-2Rβ construct where Tyr-338 is mutated to Phe. Cells expressing this mutant IL-2Rβ chain fail to induce phosphorylation of PI3K-p85α/β or activate Akt, but mediate normal IL-2-dependent proliferation and activation of JAK1 and STAT-5A/B. Microarray analyses revealed differential regulation of numerous genes compared with cells expressing a wild-type IL-2Rβ, including up-regulation of the IL-17 receptor subunit IL-17RA. Blockade of the PI3K pathway but not p70S6K led to up-regulation of IL-17RA, and constitutive Akt activation was associated with suppressed IL-17RA expression. Moreover, similar to the mutant EPO-IL-2Rβ chimera, IL-15 and IL-21 induced IL-17RA preferentially compared with IL-2, and IL-2 but not IL-15 or IL-21 mediated prolonged activation of the PI3K p85 regulatory subunit. Thus, there are intrinsic signaling differences between IL-2 and IL-15 that can be attributed to differences in activation of the PI3K pathway.

IL-17 is the defining cytokine of a newly-described “Th17” population that plays critical roles in mediating inflammation and autoimmunity. The IL-17/IL-17 receptor superfamily is the most recent class of cytokines and receptors to be described, and until recently very little was known about its function or molecular biology. However, in the last year important new insights into the composition and dynamics of the receptor complex and mechanisms of downstream signal transduction have been made, which will be reviewed here.

IL-17 is the hallmark cytokine of the newly described “Th17” lymphocyte population. The composition, subunit dynamics, and ligand contacts of the IL-17 receptor are poorly defined. We previously demonstrated that the IL-17RA subunit oligomerizes in the membrane without a ligand. In this study, computational modeling identified two fibronectin-III-like (FN) domains in IL-17RA connected by a nonstructured linker, which we predicted to mediate homotypic interactions. In yeast two-hybrid, the membrane-proximal FN domain (FN2), but not the membrane-distal domain (FN1), formed homomeric interactions. The ability of FN2 to drive ligand-independent multimerization was verified by coimmunoprecipitation and fluorescence resonance energy transfer microscopy. Thus, FN2 constitutes a “pre-ligand assembly domain” (PLAD). Further studies indicated that the FN2 linker domain contains the IL-17 binding site, which was never mapped. However, the FN1 domain is also required for high affinity interactions with IL-17. Therefore, although the PLAD is located entirely within FN2, effective ligand binding also involves contributions from the linker and FN1.

The IL-17 cytokine family plays a major role in promoting autoimmune and inflammatory diseases. The original member of this family, IL-17 (CTLA-8 or IL-17A), has been the center of intense research, whereas IL-17F is much less understood. In many respects, IL-17F functions similarly to IL-17A: both are produced by a unique subset of CD4+ T helper cells (termed TH17) and both bind a common receptor subunit (IL-17RA). Hence, IL-17F is implicated in the diseases where IL-17A clearly plays a key role. This patent describes antagonists that can directly or indirectly inhibit IL-17F signaling. The authors also report the identification of new receptor subunits, heteromeric combinations of IL-17A and IL-17F, and that IL-21 may be a stimulus for production of IL-17F. The invention of these antagonists may provide reagents to enable further research on this intriguing molecule, as well as establish a basis for treating IL-17F-mediated diseases.

IL-17 (IL-17A) has emerged as a key mediator of protection against extracellular microbes, but this cytokine also drives pathology in various autoimmune diseases. Overwhelming data in both humans and mice reveal a clear and surprisingly specific role for IL-17 in protection against the fungus Candida albicans, a commensal microbe of the human oral cavity, gastrointestinal tract, and reproductive mucosa. The IL-17 pathway regulates antifungal immunity through upregulation of proinflammatory cytokines, including IL-6, neutrophil-recruiting chemokines (e.g., CXCL1 and CXCL5), and antimicrobial peptides (e.g., defensins), which act in concert to limit fungal overgrowth. This review focuses on diseases caused by C. albicans, the role of IL-17–mediated immunity in candidiasis, and the implications for clinical therapies for both autoimmune conditions and fungal infections.

Candida is the third most common cause of bloodstream infections in hospitalized patients. Immunity to C. albicans, the most frequent species to be isolated in candidiasis, involves a well-characterized Dectin-1/caspase-associated recruitment domain adaptor 9 (CARD9)/IL-17 signaling axis. Infections caused by non-albicans Candida species are on the rise, but surprisingly little is known about immunity to these pathogens. In this study, we evaluated a systemic infection model of C. tropicalis, a clinically relevant, but poorly understood, non-albicans Candida. Mice lacking CARD9 were profoundly susceptible to C. tropicalis, displaying elevated fungal burdens in visceral organs and increased mortality compared with wild-type (WT) controls. Unlike C. albicans, IL-17 responses were induced normally in CARD9(-/-) mice following C. tropicalis infection. Moreover, there was no difference in susceptibility to C. tropicalis infection between WT and IL-23p19(-/-), IL-17RA(-/-), or Act1(-/-) mice. However, TNF-α expression was markedly impaired in CARD9(-/-) mice. Consistently, WT mice depleted of TNF-α were more susceptible to C. tropicalis, and CARD9-deficient neutrophils and monocytes failed to produce TNF-α following stimulation with C. tropicalis Ags. Both neutrophils and monocytes were necessary for defense against C. tropicalis, because their depletion in WT mice enhanced susceptibility to C. tropicalis. Disease in CARD9(-/-) mice was not due to defective neutrophil or monocyte recruitment to infected kidneys. However, TNF-α treatment of neutrophils in vitro enhanced their ability to kill C. tropicalis. Thus, protection against systemic C. tropicalis infection requires CARD9 and TNF-α, but not IL-17, signaling. Moreover, CARD9-dependent production of TNF-α enhances the candidacidal capacity of neutrophils, limiting fungal disease during disseminated C. tropicalis infection.

T helper 17 (TH17) cells are CD4+ T cells that secrete the proinflammatory cytokine interleukin-17 (IL-17) and that play a key pathogenic role in autoimmune diseases. Through inducible and tissue-specific deletion systems, we described the temporal and cell type–specific roles of the mitogen-activated protein kinase (MAPK) p38α in mediating TH17 cell–induced tissue inflammation. Inducible deletion of Mapk14 (which encodes p38α) after the onset of experimental autoimmune encephalomyelitis (EAE), a murine model for human multiple sclerosis, protected mice from inflammation. Furthermore, the severity of EAE was markedly reduced in mice with specific loss of p38α in neuroectoderm-derived cells, including astrocytes, an effect that was associated with defective production of chemokines and decreased infiltration of the target tissue by immune cells. p38α linked IL-17 receptor (IL-17R) signaling to the expression of genes encoding proinflammatory chemokines and cytokines. Mice that lacked MAPK phosphatase 1 (MKP-1), an inhibitor of p38α, had exacerbated EAE and enhanced expression of IL-17R–dependent genes. Our results suggest that the p38α–MKP-1 signaling axis links IL-17R signaling in tissue-resident cells to autoimmune inflammation dependent on infiltrating TH17 cells.

Candida albicans is a commensal fungal microbe of the human orogastrointestinal tract and skin. C. albicans causes multiple forms of disease in immunocompromised patients, including oral, vaginal, dermal and disseminated candidiasis. The cytokine IL-17 (IL-17A) and its receptor subunits, IL-17RA and IL-17RC, are required for protection to most forms of candidiasis. The importance of the IL-17R pathway has been observed not only in knockout mouse models, but also in humans with rare genetic mutations that impact generation of Th17 cells or the IL-17 signaling pathway, including Hyper-IgE Syndrome (STAT3 or TYK2 mutations) or IL17RA or ACT1 gene deficiency. The IL-17 family of cytokines is a distinct subclass of cytokines with unique structural and signaling properties. IL-17A is the best-characterized member of the IL-17 family to date, but far less is known about other IL-17-related cytokines. In this study, we sought to determine the role of a related IL-17 cytokine, IL-17C, in protection against oral, dermal and disseminated forms of C. albicans infection. IL-17C signals through a heterodimeric receptor composed of the IL-17RA and IL-17RE subunits. We observed that IL-17C mRNA was induced following oral C. albicans infection. However, mice lacking IL-17C or IL-17RE cleared C. albicans infections in the oral mucosa, skin and bloodstream at rates similar to WT littermate controls. Moreover, these mice demonstrated similar gene transcription profiles and recovery kinetics as WT animals. These findings indicate that IL-17C and IL-17RE are dispensable for immunity to the forms of candidiasis evaluated, and illustrate a surprisingly limited specificity of the IL-17 family of cytokines with respect to systemic, oral and cutaneous Candida infections.

The existence of interleukin (IL)-2 has been recognized for over 25 years, and it remains one of the most extensively studied cytokines. Here we present a broad overview of IL-2 history, functional activities, biological sources, regulation and applications to disease treatment. IL-2 exerts a wide spectrum of effects on the immune system, and it plays crucial roles in regulating both immune activation and homeostasis. Both IL-2 and its multipartite receptor are prototypical of the Type I receptor superfamily, and both have been exploited in numerous ways in the clinic. Despite the wealth of information generated about IL-2 from in vitro culture systems, in vivo mouse knockout models, and clinical trials in humans, fascinating new aspects of its functions in the immune system continue to emerge.

CARMA1 (also known as CARD11) is a scaffold molecule and contains a caspase-recruitment domain (CARD) and a membrane-associated guanylate kinase-like (MAGUK) domain. It plays an essential role in mediating CD3/CD28 costimulation-induced NF-kappaB activation. However, the molecular mechanism by which CARMA1 mediates costimulatory signals remains to be determined. Here, we show that CARMA1 is constitutively associated with the cytoplasmic membrane. This membrane association is essential for the function of CARMA1, since a mutant of CARMA1, CARMA1(L808P), that is defective in the membrane association cannot rescue CD3/CD28 costimulation-induced NF-kappaB activation in JPM50.6 CARMA1-deficient T cells. Although CD3/CD28 costimulation effectively induces the formation of the immunological synapse in CARMA1-deficient T cells, the recruitment of protein kinase C-theta (PKC-theta), Bcl10, and IkappaB kinase beta (IKKbeta) into lipid rafts of the immunological synapse is defective. Moreover, expression of wild-type CARMA1, but not CARMA1(L808P), restores the recruitment of PKC-theta, Bcl10, and IKKbeta into lipid rafts in CARMA1-deficient T cells. Consistently, expression of a mutant CARMA1, CARMA1(DeltaCD), that cannot associate with Bcl10 failed to restore CD3/CD28 costimulation-induced NF-kappaB activation in JPM50.6 cells, whereas expression of Bcl10-CARMA(DeltaCD) fusion protein effectively restored this NF-kappaB activation. Together, these results indicate that CARMA1 mediates CD3/CD28 costimulation-induced NF-kappaB activation by recruiting downstream signaling components into the immunological synapse.

Lymphocyte populations in the immune system are maintained by a well-organized balance between cellular proliferation, cellular survival and programmed cell death (apoptosis). One of the primary functions of many cytokines is to coordinate these processes. In particular, the interleukin (IL)-2 family of cytokines, which consists of six cytokines (IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21) that all share a common receptor subunit (gammac), plays a major role in promoting and maintaining T lymphocyte populations. The details of the molecular signaling pathways mediated by these cytokines have not been fully elucidated. However, the three major pathways clearly involved include the JAK/STAT, MAPK and phosphatidylinositol 3-kinase (P13K) pathways. The details of these pathways as they apply to the IL-2 family of cytokines is discussed, with a focus on their roles in proliferation and survival signaling.

The interleukin-2 receptor (IL-2R) is composed of one affinity-modulating subunit (IL-2Ralpha) and two essential signaling subunits (IL-2Rbeta and gammac). Although most known signaling events are mediated through tyrosine residues located within IL-2Rbeta, no functions have yet been ascribed to gammac tyrosine residues. In this study, we describe a role for gammac tyrosines in anti-apoptotic signal transduction. We have shown previously that a tyrosine-deficient IL-2Rbeta chain paired with wild type gammac stimulated enhancement of bcl-2 mRNA in IL-2-dependent T cells, but it was not determined which region of the IL-2R or which pathway was activated to direct this signaling response. Here we show that up-regulation of Bcl-2 by an IL-2R lacking IL-2Rbeta tyrosine residues leads to increased cell survival after cytokine deprivation; strikingly, this survival signal does not occur in the absence of gammac tyrosine residues. These gammac-dependent signals are revealed only in the absence of IL-2Rbeta tyrosines, indicating that the IL-2R engages at least two distinct signaling pathways to regulate apoptosis and Bcl-2 expression. Mechanistically, the gammac-dependent signal requires activation of Janus kinases 1 and 3 and is sensitive to wortmannin, implicating phosphatidylinositol 3-kinase. Consistent with involvement of phosphatidylinositol 3-kinase, Akt can be activated via tyrosine residues on gammac. Thus, gammac mediates an anti-apoptotic signaling pathway through Akt which cooperates with signals from its partner chain, IL-2Rbeta.

Disseminated candidiasis is the fourth most common hospital acquired infection and is associated with a 40–60% mortality rate. During disseminated candidiasis, Candida albicans form invasive hyphae that damage kidney, leading to renal failure. Treatment of candidiasis is hindered by drug toxicity, the development of antifungal drug resistance and lack of vaccines against fungal pathogens. Previous studies have identified a key role of interleukin-17 (IL-17) in controlling systemic infection. The mechanisms of IL-17-mediated renal immunity have so far been assumed to occur solely through the regulation of antimicrobial mechanisms. Here, we discovered an unanticipated role for IL-17 in inducing the Kallikrein (Klk)-Kinin System (KKS) in the infected kidney, and we show that the KKS provides significant renal protection in candidiasis. Consequently, overexpression of Klk1 or treatment with bradykinin rescued IL-17RA−/− mice from candidiasis. Therapeutic manipulation of IL-17-KKS pathways restored renal function and prolonged survival by preventing apoptosis of renal cells following disseminated infection. Moreover, combining a minimally effective dose of fluconazole with bradykinin strikingly improved survival compared to either drug alone. These findings have potential translational significance, as agonists of the KKS are in routine clinical use. Therefore, these results not only identify downstream mediators that could serve as novel drug targets, but could possibly be used to guide decisions on whether targeting these mediators could be a useful therapeutic option in conjunction with current antifungal drugs.

IL-17 is a pro-inflammatory cytokine that drives autoimmune diseases and promotes immunity to fungi. IL-17 binding to the IL-17 receptor initiates recruitment of adaptor proteins that activate NF-κB. IL-17 also induces expression of A20, a feedback inhibitor that restricts NF-κB activation and limits IL-17-dependent inflammation. ABIN1 is an A20 binding protein that has been implicated in autoimmunity in human genetic studies, so we hypothesized that ABIN1 might also play a role in restricting IL-17-dependent signaling. Indeed, ABIN1 inhibited the IL-17 pathway based on RNA silencing analyses. Specifically, the expression of the IL-17-dependent gene lipocalin 2 (Lcn2) increased at both basal levels and after stimulation upon ABIN1 knockdown. Consistently, reconstitution of ABIN1-deficient fibroblasts with ABIN1 also decreased both Lcn2 expression and promoter activation. However, while ABIN1 mRNA increased after IL-17A stimulation, ABIN1 protein progressively decreased following its inducible phosphorylation. Although feedback loops are commonly seen in signaling pathways, differential regulation of mRNA and protein of the same gene are not as common. Thus, we now propose that ABIN1 serves as a constitutive inhibitor of NF-κB signaling. While ABIN1 mRNA is induced by NF-κB at the mRNA level, removal of the ABIN1 protein is required to initiate signal transduction. Unexpectedly, we discovered that A20 is dispensable for both IL-17-mediated degradation of ABIN1 and for ABIN1-dependent inhibition of IL-17 signaling. Altogether, these findings suggest that ABIN1 is a tonic NF-κB inhibitor, and that IL-17 signaling induces its degradation independently of A20 to enhance expression of NF-κB-dependent genes such as Lcn2.

Candida albicans is normally a commensal fungus of the human mucosae and skin, but it causes life-threatening systemic infections in hospital settings in the face of predisposing conditions, such as indwelling catheters, abdominal surgery, or antibiotic use. Immunity to C. albicans involves various immune parameters, but the cytokine interleukin-17A (IL-17A) (also known as IL-17) has emerged as a centrally important mediator of immune defense against both mucosal and systemic candidiasis. Conversely, IL-17A has been suggested to enhance the virulence of C. albicans , indicating that it may exert detrimental effects on pathogenesis. In this study, we hypothesized that a C. albicans strain expressing IL-17A would exhibit reduced virulence in vivo . To that end, we created a Candida -optimized expression cassette encoding murine IL-17A, which was transformed into the DAY286 strain of C. albicans . Candida -derived IL-17A was indistinguishable from murine IL-17A in terms of biological activity and detection in standard enzyme-linked immunosorbent assays (ELISAs). Expression of IL-17A did not negatively impact the growth of these strains in vitro . Moreover, the IL-17A-expressing C. albicans strains showed significantly reduced pathogenicity in a systemic model of Candida infection, mainly evident during the early stages of disease. Collectively, these findings suggest that IL-17A mitigates the virulence of C. albicans .

Human fungal pathogens are responsible for more than a million life-threatening infections annually, which can be associated with mortality rates reaching 95% (Brown et al. 2012). Fungal infections are difficult to treat and control because of rising problems of antifungal drug resistance and the lack of diagnostics, novel antifungal drugs, and vaccines. FortW1ately, the majority of pathogenic fungi are opportunistic pathogens (Table 1) and, as such, do not normally cause disease unless there are alternations in immune defense. The use of immunosuppressive drugs, the human immunodeficiency virus (HIV) epidemic, and modern clinical interventions result in such alternations and have contributed substantially to the recent increases of systemically infected patients. Fungi also cause nonlethal skin and mucosal infections that are equally difficult to treat and often recurring. Table 1 Species of pathogenic fungi that cause disease in humans, the barrier tissue breached and organs infected, and the PRRs needed for protective immunity The innate immune system is the first line of defense against pathogens and broadly protects against invading microorganisms. Genetically inherited receptors, called pattern recognition receptors (PRRs), are used by innate cells for recognition of conserved pathogen-associated molecular patterns. Signaling downstream from PRRs activates cellular responses and killing mechanisms, and also helps initiate and shape adaptive immune responses. Adaptive immunity, unlike innate, is activated by specific antigens recognized by noninherited T-cell/B-cell receptors and has a faster response time with each subsequent challenge (immune memory) (Wuthrich et al. 2012). The vast predominance of fungi in the environment results in continual human exposure. It is estimated that we inhale several hundred Aspergillus fumigatus spores a day (Rivera et al. 2011), and many humans are also colonized with commensal fungi (e.g., Candida albicans) (Iliev et al. 2012). For this reason, our innate system is equipped to recognize fungal particles and maintain commensal relationships, but also destroy the pathogenic fungi that we are exposed to. This review will outline the cells and recognition receptors in innate antifungal immunity, with a special focus on IL-17 defenses and inflammasomes, both recently identified as major players in antifungal defense.

In recent years, proinflammatory cytokines in the nephritic kidney appear to contribute to the pathogenesis of AGN. The complex inflammatory cytokine network that drives renal pathology is poorly understood. IL-17, the signature cytokine of Th17 cells, which promotes autoimmune pathology in a variety of settings, is beginning to be identified in acute and chronic kidney diseases as well. However, the role of IL-17-mediated renal damage in the nephritic kidney has not been elucidated. Here, with the use of a murine model of experimental AGN, we showed that IL-17RA signaling is critical for the development of renal pathology. Despite normal systemic autoantibody response and glomerular immune-complex deposition, IL-17RA−/− mice exhibit a diminished influx of inflammatory cells and kidney-specific expression of IL-17 target genes correlating with disease resistance in AGN. IL-17 enhanced the production of proinflammatory cytokines and chemokines from tECs. Finally, we were able to show that neutralization of IL-17A ameliorated renal pathology in WT mice following AGN. These results clearly demonstrated that IL-17RA signaling significantly contributes to renal tissue injury in experimental AGN and suggest that blocking IL-17RA may be a promising therapeutic strategy for the treatment of proliferative and crescentic glomerulonephritis.

Multiple forms of candidiasis are clinically important in humans. Established murine models of disseminated, oropharyngeal, vaginal, and cutaneous candidiasis caused by Candida albicans are described in this unit. Detailed materials and methods for C. albicans growth and detection are also described.

Oropharyngeal candidiasis (OPC; thrush) is an opportunistic fungal infection caused by the commensal microbe Candida albicans . Immunity to OPC is strongly dependent on CD4 + T cells, particularly those of the Th17 subset. Interleukin-17 (IL-17) deficiency in mice or humans leads to chronic mucocutaneous candidiasis, but the specific downstream mechanisms of IL-17-mediated host defense remain unclear. Lipocalin 2 (Lcn2; 24p3; neutrophil gelatinase-associated lipocalin [NGAL]) is an antimicrobial host defense factor produced in response to inflammatory cytokines, particularly IL-17. Lcn2 plays a key role in preventing iron acquisition by bacteria that use catecholate-type siderophores, and lipocalin 2 −/− mice are highly susceptible to infection by Escherichia coli and Klebsiella pneumoniae . The role of Lcn2 in mediating immunity to fungi is poorly defined. Accordingly, in this study, we evaluated the role of Lcn2 in immunity to oral infection with C. albicans . Lcn2 is strongly upregulated following oral infection with C. albicans , and its expression is almost entirely abrogated in mice with defective IL-17 signaling (IL-17RA −/− or Act1 −/− mice). However, Lcn2 −/− mice were completely resistant to OPC, comparably to wild-type (WT) mice. Moreover, Lcn2 deficiency mediated protection from OPC induced by steroid immunosuppression. Therefore, despite its potent regulation during C. albicans infection, Lcn2 is not required for immunity to mucosal candidiasis.

Erythropoietin (EPO) and its receptor (EPOR) are required for the development of mature erythrocytes. After binding of ligand, the EPOR activates a variety of signaling pathways that ultimately control cellular proliferation, survival, and specific gene expression. Although erythroid progenitors appear to be the principal EPO-responsive cell type in vivo due to the restricted expression of the EPOR, many growth factor–dependent cell lines expressing the EPOR can respond to EPO by activating many or all of these pathways. In the present study, we have identified a cellular context (the interleukin-2 [IL-2]–dependent HT-2 line) in which the EPO stimulation of the EPOR fails to support cellular proliferation, STAT-5 induction, or MAPK activation, despite efficient phosphorylation of the EPOR and JAK2 and inhibition of apoptosis after withdrawal of IL-2. Interestingly, when we fused HT-2 cells expressing the EPOR with Ba/F3 cells in a complementation assay, the resulting hybridomas proliferated and potently activated STAT-5 and MAPK in response to EPO. These data indicate that an unidentified cellular factor is needed to mediate signaling by the EPOR. Moreover, Ba/F3 cells apparently express this factor(s) and somatic fusions can, therefore, confer EPO-responsiveness to HT-2 cells that lack this factor.

The cytokine IL-17 came into prominence a decade ago with the discovery of Th17 cells. IL-17 signals through its receptor to control immune responses to extracellular pathogens, particularly fungi. Conversely, IL-17 activates pathology in many autoimmune and infectious settings. Therefore, constraint of the IL-17 receptor pathway is an essential facet of its regulation. To date, our understanding of IL-17 signaling checkpoints remains quite limited, but in recent years a number of negative regulators have been identified. These inhibitors block IL-17 activation at multiple levels, although the integration of these inhibitors remains poorly understood. This talk will discuss emerging work defining several novel feedback inhibitors that restrict the downstream IL-17 signaling pathway and their molecular mechanisms of action.

“For every action, there is an equal and opposite reaction.” This axiom is true not only for physical processes, but also for biological activation events, where inhibitory mechanisms are in place to reestablish equilibrium. Nowhere is this more evident than during acute inflammation, where cytokines or pathogen-associated stimuli ramp up a rapid and powerful inflammatory cascade, followed by an equally important phase of resolution. Keeping inflammation in check avoids collateral tissue damage, ultimately preventing autoimmune and hyperproliferative disorders. A20 is a vital inhibitor of inflammation encoded by the gene TNFα-induced protein 3 (TNFAIP3). The TNFAIP3 locus is linked to autoimmune diseases and B-cell lymphomas.1 A20 is a deubiquitinating enzyme that reverses Lysine (K)63-linked ubiquitination. A20 was initially described as a negative feedback inhibitor of TNFα-induced signaling, where it acts principally by targeting RIP1, and thereby terminating activation of the NFκB transcription factor. A20 also inhibits the TLR, IL-1R, and Nod-like receptor pathways, where it targets other signaling intermediates such as TRAF6, NEMO, and Ubc13 (Fig. 1).2 Mathematical modeling of the TNFα pathway led to the insight that A20 restricts activation of the second, not the first phase of classical NFκB activation. Moreover, due to its long protein half-life, steady-state levels of A20 dictate the magnitude of inflammatory signaling. Notably, induction of A20 by any stimulus is predicted to exert a regulatory impact on the global response; thus, A20 is considered a “rheostat” that maintains equilibrium following NFκB activation.3 Figure 1. Schematic diagram depicting A20-mediated regulation of inflammatory pathways. The model represents the two phases of feedback inhibition of inflammation with A20 (tnfaip3) as a central player. In the first phase, several stimuli such as TNFα, ... Confirming the importance of A20 in restricting inflammation, A20−/− mice die prematurely of multi-organ inflammation and cachexia. Surprisingly, despite the prominent role of A20 in tempering TNFα signaling, inflammation in A20−/− mice was not alleviated by crossing to mice lacking the TNF receptor. Subsequent in vivo studies revealed that the baseline inflammation in A20-deficient mice is due to signals from commensal microbiota via TLR/MyD88 signaling. A number of subsequent studies examined cell type-specific effects of A20 using conditionally targeted mice, demonstrating important activities for this protein in numerous immune subtypes.1 In recent years IL-17 has come to the limelight due the recognition of Th17 cells and their connection to autoimmune disease. Clinically, antibodies against IL-17 or its receptor are showing promise in treating a variety of other conditions.4 The IL-17R belongs to a distinct subfamily of receptors, with many unique structural and functional properties. Nonetheless, the IL-17 signaling pathway shares many downstream adaptors and transcription factors with the TNFα, IL-1, and TLR pathways.5 Given these commonalities, we predicted that A20 might also serve as an inhibitor of IL-17 signaling. Indeed, IL-17 induced expression of the TNFAIP3 gene in an NFκB-dependent manner. A20 deficiency upon siRNA silencing or in knockout cells led to enhanced IL-17-dependent expression of inflammatory genes. Mechanistically, A20 binds to and dequbitinates TRAF6, ultimately inactivating IL-17-mediated NF-κB and MAPK (particularly JNK) pathways.6 Although the mechanism used by A20 to inhibit IL-17 signaling has parallels with the IL-1R pathway, there are striking differences. In other pathways, A20 interacts with accessory proteins such as Tax1bp1 and Itch to efficiently hydrolyze K63-linked substrates. Deletion of these proteins results in a phenotype similar to the lack of A20, though less severe.2 Unexpectedly, Tax1bp1 and Itch were dispensable for A20s activity in IL-17 signaling, as siRNA-mediated silencing of Tax1bp1 or Itch did not affect IL-17-dependent gene expression.7 This finding raised the possibility that A20 either functions solo or uses alternate adaptor molecules. Accordingly, we found that A20 associates with AnapC5 (APC5), a subunit of a multi-protein E3 ubiquitin ligase complex known as the Anaphase Promoting Complex (APC). APC subunits have not been previously linked with the regulation of inflammatory signaling, suggesting a novel role for AnapC5 outside of regulating the cell cycle (Fig. 1). In analyzing how A20 is recruited to the IL-17R signaling complex, we found that A20 associates directly with the IL-17RA subunit.6 Notably, A20 binds to a C-terminal domain of IL-17RA that was previously shown to downmodulate IL-17 signaling.8 To our knowledge, this is the first demonstration that A20 associates with an inducing receptor. A20 interaction with receptors may also occur in other systems, potentially serving as a mechanism to recruit this inhibitor after a signaling event is triggered. These discoveries about A20 function in the context of IL-17 suggest there are distinct mechanisms by which A20 restricts inflammatory pathways. It will be key to define the contribution of A20 in regulating IL-17 in vivo. As noted, SNPs in the TNFAIP3 gene locus are associated with autoimmune diseases that are linked to IL-17 activity. Therefore, it would not be surprising if patients with TNFAIP3 polymorphisms are more likely to be responsive to anti-IL-17 therapy, as has been demonstrated for TNF inhibitors. To date, therapeutic strategies to block IL-17 have mainly focused on biologic drugs that target Th17 cells (anti-IL-6 or anti-IL-12/23) or directly neutralize this cytokine (anti-IL-17, anti-IL-17RA). Development of small-molecule inhibitors will be aided by a better understanding of how IL-17 pathways are regulated in nature.

The proinflammatory cytokine interleukin-17 (IL-17) is the signature cytokine of the T helper 17 (TH17) subset of CD4(+) T cells, and antibodies targeting IL-17 or the IL-17 receptor (IL-17R) show clinical efficacy in several autoimmune diseases. Although important for protective immunity against microorganisms, IL-17 causes collateral damage in inflammatory settings. TNFAIP3 encodes the deubiquitinase A20 and is genetically linked to numerous autoimmune syndromes. A20, a potent inhibitor of tumor necrosis factor-α signaling, removes ubiquitin from signaling intermediates upstream of nuclear factor κB (NF-κB), thereby dampening NF-κB-mediated inflammation. We demonstrated that IL-17 stimulates TNFAIP3 expression. Enhanced IL-17-mediated induction of genes encoding proinflammatory factors, including IL-6 and various chemokines, occurred upon knockdown of A20 with short inhibitory RNA or in A20(-/-) cells. A20 associated with the E3 ubiquitin ligase TRAF6 (tumor necrosis factor receptor-associated factor 6) in an IL-17-dependent manner and restricted the IL-17-dependent activation of NF-κB and mitogen-activated protein kinases. A20 interacted directly with the distal domain of IL-17RA, a previously defined inhibitory domain. Together, these data describe a mechanism of restraining IL-17 signaling and reveal an aspect of A20 activity that may help to explain its role in autoimmunity in humans.

Candida albicans remains the fungus most frequently associated with nosocomial bloodstream infection. In disseminated candidiasis, the role of Foxp3(+) regulatory T (Treg) cells remains largely unexplored. Our aims were to characterize Foxp3(+) Treg-cell activation in a murine intravenous challenge model of disseminated C. albicans infection, and determine the contribution to disease. Flow cytometric analyses demonstrated that C. albicans infection drove in vivo expansion of a splenic CD4(+) Foxp3(+) population that correlated positively with fungal burden. Depletion from Foxp3(hCD2) reporter mice in vivo confirmed that Foxp3(+) cells exacerbated fungal burden and inflammatory renal disease. The CD4(+) Foxp3(+) population expanded further after in vitro stimulation with C. albicans antigens (Ags), and included at least three cell types. These arose from proliferation of the natural Treg-cell subset, together with conversion of Foxp3(-) cells to the induced Treg-cell form, and to a cell type sharing effector Th17-cell characteristics, expressing ROR-γt, and secreting IL-17A. The expanded Foxp3(+) T cells inhibited Th1 and Th2 responses, but enhanced Th17-cell responses to C. albicans Ags in vitro, and in vivo depletion confirmed their ability to enhance the Th17-cell response. These data lead to a model for disseminated candidiasis whereby expansion of Foxp3(+) T cells promotes Th17-cell responses that drive pathology.

The specific signal transduction function of the gamma c subunit in the interleukin (IL) 2, IL-4, IL-7, IL-9, and IL-15 receptor complexes remains undefined. The present structure-function analyses demonstrated that the entire cytoplasmic tail of gamma c could be functionally replaced in the IL-2 receptor (IL-2R) signaling complex by a severely truncated erythropoietin receptor cytoplasmic domain lacking tyrosine residues. Heterodimerization of IL-2R beta with either gamma c or the truncated erythropoietin receptor chain led to an array of specific signals normally derived from the native IL-2R despite the substitution of Janus kinase JAK2 for JAK3 in the receptor complex. These findings thus suggest a model in which the gamma c subunit serves as a common and generic "trigger" chain by providing a nonspecific Janus kinase for signaling program initiation, while signal specificity is determined by the unique "driver" subunit in each of the gamma c- containing receptor complexes. Furthermore, these results may have important functional implications for the asymmetric design of many cytokine receptor complexes and the evolutionary design of receptor subfamilies that share common trigger or driver subunits.

The incidence of life-threatening disseminated Candida albicans infections is increasing in hospitalized patients, with fatalities as high as 60%. Death from disseminated candidiasis in a significant percentage of cases is due to fungal invasion of the kidney, leading to renal failure. Treatment of candidiasis is hampered by drug toxicity, the emergence of antifungal drug resistance and lack of vaccines against fungal pathogens. IL-17 is a key mediator of defense against candidiasis. The underlying mechanisms of IL-17-mediated renal immunity have so far been assumed to occur solely through the regulation of antimicrobial mechanisms, particularly activation of neutrophils. Here, we identify an unexpected role for IL-17 in inducing the Kallikrein (Klk)-Kinin System (KKS) in C. albicans-infected kidney, and we show that the KKS provides significant renal protection in candidiasis. Microarray data indicated that Klk1 was upregulated in infected kidney in an IL-17-dependent manner. Overexpression of Klk1 or treatment with bradykinin rescued IL-17RA-/- mice from candidiasis. Therapeutic manipulation of IL-17-KKS pathways restored renal function and prolonged survival by preventing apoptosis of renal cells following C. albicans infection. Furthermore, combining a minimally effective dose of fluconazole with bradykinin markedly improved survival compared to either drug alone. These results indicate that IL-17 not only limits fungal growth in the kidney, but also prevents renal tissue damage and preserves kidney function during disseminated candidiasis through the KKS. Since drugs targeting the KKS are approved clinically, these findings offer potential avenues for the treatment of this fatal nosocomial infection., Author Summary Candida albicans is the causative agent of oropharyngeal candidiasis (OPC, thrush), dermal and vaginal candidiasis. However, the most severe C. albicans-induced disease is disseminated candidiasis, a frequent nosocomial infection associated with a high mortality rate. During disseminated candidiasis, C. albicans form invasive hyphae that damage target organs, particularly kidney and liver. Previous studies have identified an essential role of interleukin-17 (IL-17) in controlling systemic infection through regulation of neutrophils. We show here for the first time that IL-17 also regulates the renal protective Kallikrein-kinin system (KKS). Our discovery of a connection between IL-17 and the KKS suggests a new, previously unanticipated avenue for the treatment of renal damage in disseminated candidiasis. These findings have potential translational significance, as agonists of the KKS are in routine clinical use. Therefore, these results not only identify downstream mediators that could serve as novel drug targets, but could possibly be used to guide decisions on whether targeting these mediators could be a useful therapeutic option in conjunction with current antifungal therapies.

Herpesvirus Saimiri gene 13 (HVS13) exhibits 57% identity with the predicted sequence of a T cell-derived molecule termed CTLA8. Recombinant HVS13 and CTLA8 stimulate transcriptional factor NF-kappaB activity and Interleukin-6 (IL-6) secretion in fibroblasts, and costimulate T cell proliferation. An HVS13.Fc fusion protein was used to isolate a cDNA encoding a novel receptor that also binds CTLA8. This receptor is unrelated to previously identified cytokine receptor families. A recombinant soluble receptor inhibited T cell proliferation and IL-2 production induced by PHA, concanavalin A (conA), and anti-TCR MAb. These results define CTLA8 and HVS13 as novel cytokines that bind to a novel cytokine receptor. We propose to call these molecules IL-17, vIL-17, and IL-17R, respectively.

The discovery of the Th17 lineage in 2005 triggered a major change in how immunity to infectious diseases is viewed. Fungal infections, in particular, have long been a relatively understudied area of investigation in terms of the host immune response. Candida albicans is a commensal yeast that colonizes mucosal sites and skin. In healthy individuals, it is non-pathogenic, but in conditions of immune deficiency, this organism can cause a variety of infections associated with considerable morbidity. Candida can also cause disseminated infections that have a high mortality rate and are a major clinical problem in hospital settings. Although immunity to Candida albicans was long considered to be mediated by Th1 cells, new data in both rodent models and in humans have revealed an essential role for the Th17 lineage, and in particular its signature cytokine IL-17.

The IL-17/IL-17 receptor family is the newest and least understood of the cytokine subclasses. Composed of ligands IL-17A-IL-17F and receptors IL-17RA-IL-17RE, these cytokines have many unique structural and functional features. Since the discovery of the 'Th17' subset in 2005, particular attention has been paid to IL-17A and IL-17F and their cognate receptors. To date, far less is known about the rest of the family. This review discusses recent advances in the field, with an emphasis on IL-17A biology.

The IL-17/IL-17 receptor family is the newest family of cytokine ­systems to be defined. Although considerable progress has been made in recent years to decipher the structure-function relationships and downstream signaling events mediated by this family, there are still many gaps in our understanding of this area. This chapter will provide an overview of the molecular biology of the IL-17R ­family, with a major focus on the IL-17A/F and IL-17RA/IL-17RC families. Some discussion of the IL-17E/IL-25 and IL-17RB system and the SEF/IL-17RD system will also be reviewed.

Candida albicans causes mucosal and disseminated candidiasis, which represent serious problems for the rapidly expanding immunocompromised population. Until recently, Th1-mediated immunity was thought to confer the primary protection, particularly for oral candidiasis. However, emerging data indicate that the newly-defined Th17 compartment appears to play the predominant role in mucosal candidiasis.

BackgroundChronic granulomatous disease (CGD), an inherited disorder of the NADPH oxidase in which phagocytes are defective in generating superoxide anion and downstream reactive oxidant intermediates (ROIs), is characterized by recurrent bacterial and fungal infections and by excessive inflammation (e.g., inflammatory bowel disease). The mechanisms by which NADPH oxidase regulates inflammation are not well understood.Methodology/principal findingsWe found that NADPH oxidase restrains inflammation by modulating redox-sensitive innate immune pathways. When challenged with either intratracheal zymosan or LPS, NADPH oxidase-deficient p47(phox-/-) mice and gp91(phox)-deficient mice developed exaggerated and progressive lung inflammation, augmented NF-kappaB activation, and elevated downstream pro-inflammatory cytokines (TNF-alpha, IL-17, and G-CSF) compared to wildtype mice. Replacement of functional NADPH oxidase in bone marrow-derived cells restored the normal lung inflammatory response. Studies in vivo and in isolated macrophages demonstrated that in the absence of functional NADPH oxidase, zymosan failed to activate Nrf2, a key redox-sensitive anti-inflammatory regulator. The triterpenoid, CDDO-Im, activated Nrf2 independently of NADPH oxidase and reduced zymosan-induced lung inflammation in CGD mice. Consistent with these findings, zymosan-treated peripheral blood mononuclear cells from X-linked CGD patients showed impaired Nrf2 activity and increased NF-kappaB activation.Conclusions/significanceThese studies support a model in which NADPH oxidase-dependent, redox-mediated signaling is critical for termination of lung inflammation and suggest new potential therapeutic targets for CGD.

Interleukin (IL)-17 (also known as IL-17A), the signature cytokine of the newly described T helper 17 (Th17) cell population, has been implicated in the pathogenesis of numerous autoimmune diseases including rheumatoid arthritis. IL-17 is the founding member of a new subclass of cytokines that have highly proinflammatory properties. Studies in rodents and mammalian cell culture systems, as well as clinical settings, support a role for IL-17 in promoting rheumatoid arthritis. This article discusses the history of the discovery of Th17 cells, the potential mechanisms of action of IL-17 in autoimmunity, and perspectives for IL-17-targeted cytokine therapy.

TLR stimulation triggers a signaling pathway via MyD88 and IL-1R-associated kinase 4 that is essential for proinflammatory cytokine induction. Although NF-κB has been shown to be one of the key transcriptional regulators of these cytokines, evidence suggests that other factors may also be important. In this study, we showed that MyD88-deficient macrophages have defective c-Rel activation, which has been linked to IL-12p40 induction, but not IL-6 or TNF-α. We also investigated other transcription factors and showed that C/EBPβ and C/EBPδ expression was limited in MyD88- or IL-1R-associated kinase 4-deficient macrophages treated with LPS. Importantly, the absence of both C/EBPβ and C/EBPδ resulted in the impaired induction of proinflammatory cytokines stimulated by several TLR ligands. Our results identify c-Rel and C/EBPβ/δ as important transcription factors in a MyD88-dependent pathway that regulate the induction of proinflammatory cytokines.

Interleukin-17 (IL-17), the hallmark cytokine of T helper 17 (T H 17) cells, signals through a distinct receptor subclass, yet little is known about the mechanisms involved. IL-17 activates the expression of target genes through the actions of the transcription factors nuclear factor κB (NF-κB), CAAT enhancer binding protein δ (C/EBPδ), and C/EBPβ. The adaptor proteins tumor necrosis factor receptor–associated factor 6 (TRAF6) and Act1 are upstream of NF-κB and C/EBPδ, but the regulation of C/EBPβ remains undefined. Here, we show that IL-17 signaling led to phosphorylation of two sites in the regulatory 2 domain of C/EBPβ in a sequential, interdependent fashion. The first was rapid and dependent on extracellular signal–regulated kinase (ERK), whereas the second was dependent on the activity of glycogen synthase kinase 3β (GSK-3β). These pathways were mediated by distinct subdomains within IL-17 receptor A (IL-17RA). Whereas phosphorylation of threonine 188 (Thr 188 ) was mediated by the previously identified SEF/IL-17R homology domain–Toll-IL-1R–like loop (SEFIR-TILL), phosphorylation of Thr 179 occurred through a newly characterized motif located in the distal tail of IL-17RA. Phosphorylated C/EBPβ mediated a negative signal, because blocking ERK and GSK-3β increased expression of IL-17 target genes and a C/EBPβ-Thr 188 mutant enhanced activation of a C/EBP-dependent reporter. Overexpression of GSK-3β inhibited IL-17–induced activation of a C/EBP-dependent reporter, and Thr 179 of C/EBPβ was not phosphorylated in GSK-3β–deficient cells. Thus, IL-17 triggered the dual phosphorylation of C/EBPβ, which inhibited the expression of proinflammatory genes. This detailed dissection is the first for the IL-17–mediated C/EBP pathway and the first known example of a negative signal mediated by IL-17RA.

The commensal fungus Candida albicans causes oropharyngeal candidiasis (OPC; thrush) in settings of immunodeficiency. Although disseminated, vaginal, and oral candidiasis are all caused by C. albicans species, host defense against C. albicans varies by anatomical location. T helper 1 (Th1) cells have long been implicated in defense against candidiasis, whereas the role of Th17 cells remains controversial. IL-17 mediates inflammatory pathology in a gastric model of mucosal candidiasis, but is host protective in disseminated disease. Here, we directly compared Th1 and Th17 function in a model of OPC. Th17-deficient (IL-23p19(-/-)) and IL-17R-deficient (IL-17RA(-/-)) mice experienced severe OPC, whereas Th1-deficient (IL-12p35(-/-)) mice showed low fungal burdens and no overt disease. Neutrophil recruitment was impaired in IL-23p19(-/-) and IL-17RA(-/-), but not IL-12(-/-), mice, and TCR-alphabeta cells were more important than TCR-gammadelta cells. Surprisingly, mice deficient in the Th17 cytokine IL-22 were only mildly susceptible to OPC, indicating that IL-17 rather than IL-22 is vital in defense against oral candidiasis. Gene profiling of oral mucosal tissue showed strong induction of Th17 signature genes, including CXC chemokines and beta defensin-3. Saliva from Th17-deficient, but not Th1-deficient, mice exhibited reduced candidacidal activity. Thus, the Th17 lineage, acting largely through IL-17, confers the dominant response to oral candidiasis through neutrophils and antimicrobial factors.

Interleukin-17 (IL-17) is a proinflammatory cytokine secreted by the newly described CD4+Th17 subset, which is distinct from classic Th1 and Th2 lineages. IL-17 contributes to bone destruction in rheumatoid arthritis but is essential in host defense against pathogens that are susceptible to neutrophils. Periodontal disease (PD) is a chronic inflammatory condition initiated by anaerobic oral pathogens such asPorphyromonas gingivalis, and it is characterized by host-mediated alveolar bone destruction due primarily to the immune response. The role of IL-17 in PD is controversial. Whereas elevated IL-17 levels have been found in humans with severe PD, we recently reported that female C57BL/6J mice lacking the IL-17 receptor (IL-17RAKO) are significantly more susceptible to PD bone loss due to defects in the chemokine-neutrophil axis (J. J. Yu, M. J. Ruddy, G. C. Wong, C. Sfintescu, P. J. Baker, J. B. Smith, R. T. Evans, and S. L. Gaffen, Blood109:3794-3802, 2007). Since different mouse strains exhibit differences in susceptibility to PD as well as Th1/Th2 cell skewing, we crossed the IL-17RA gene knockout onto the BALB/c background and observed a similar enhancement in alveolar bone loss followingP. gingivalisinfection. Unexpectedly, in both strains IL-17RAKOfemale mice were much more susceptible to PD bone loss than males. Moreover, female BALB/c-IL-17RAKOmice were defective in producing anti-P. gingivalisimmunoglobulin G and the chemokines KC/Groα and MIP-2. In contrast, male mice produced normal levels of chemokines and anti-P. gingivalisantibodies, but they were defective in granulocyte colony-stimulating factor upregulation. This study demonstrates a gender-dependent effect of IL-17 signaling and indicates that gender differences should be taken into account in the preclinical and clinical safety testing of anti-IL-17 biologic therapies.

Interleukin-17 (IL-17A) is a pro-inflammatory cytokine that is primarily secreted from T lymphocytes, mediators of adaptive immunity. Recently, IL-17 was shown to be the defining cytokine of a new T helper subset termed "Th17." Discovery of the Th17 population was a groundbreaking discovery that has triggered major revisions of the prevailing paradigms in T cell biology. Although produced by T cells, IL-17 promotes expansion and recruitment of innate immune cells such as neutrophils, and also cooperates with TLR ligands, IL-1 beta, and TNF alpha to enhance inflammatory reactions and stimulate production of beta-defensins and other antimicrobial peptides. Its receptor, IL-17RA, is ubiquitously expressed and shares many features with classical innate immune receptors such as shared intracellular tail motifs and convergence on common inflammatory transcription pathways. The role of IL-17 in periodontal disease is still uncertain, since IL-17 has been shown to promote bone destruction in arthritis, but is nonetheless essential to protect the host from pathogens, including periodontopathic organisms. Recent evidence has shown that Th17 cells are more osteoclastogenic than other T helper subsets such as Th1 or Th2. Ablation of IL-17 signaling prior to onset of infection with Porphyromonas gingivalis increases susceptibility to periodontal bone loss, but this finding does not rule out the efficacy of therapeutic inhibition of IL-17 after onset of severe disease. IL-17 sits at the center of many complex diseases that integrate innate and adaptive immune mechanisms and requires careful study to maximize host protective effects and minimize host deleterious effects.

Chronic diseases, such as periodontal disease (PD) and rheumatoid arthritis (RA), are characterized by a robust immune response resulting in unresolved inflammation. Inflammation is mediated by proinflammatory cytokines; recently, a novel subset of T-helper (Th) cells was identified that plays a crucial role in inflammation and autoimmune disease. This population secretes several proinflammatory cytokines, including the novel cytokine interleukin (IL)-17, and, hence, has been termed "Th17." Inflammatory cytokines are implicated in the progression of localized chronic infections, such as PD, and in serious systemic pathologies, such as diabetes, chronic obstructive pulmonary disease, and cardiovascular disease. IL-17 mediates inflammation through a receptor (IL-17R) composed of two subunits, IL-17RA and IL-17RC. Drugs that antagonize inflammatory cytokines are used therapeutically to downregulate immune-mediated pathology in conditions such as RA, although not all patients respond well to this approach. Therefore, identification of potential novel therapeutic targets, such as the IL-17 signaling complex, may be clinically relevant for mitigating inflammatory pathology. However, the manner in which such a therapeutic may influence the onset and progression of PD is poorly understood. Therapeutics that antagonize inflammatory cytokines ameliorate inflammation and bone loss and may have broader implications for individuals with systemic diseases in which inflammation and autoimmunity predominate.

IL-17 is the founding member of a novel family of proinflammatory cytokines that defines a new class of CD4 + effector T cells, termed “Th17.” Mounting evidence suggests that IL-17 and Th17 cells cause pathology in autoimmunity, but little is known about mechanisms of IL-17RA signaling. IL-17 through its receptor (IL-17RA) activates genes typical of innate immune cytokines, such as TNFα and IL-1β, despite minimal sequence similarity in their respective receptors. A previous bioinformatics study predicted a subdomain in IL-17-family receptors with homology to a Toll/IL-1R (TIR) domain, termed the “SEFIR domain.” However, the SEFIR domain lacks motifs critical for bona fide TIR domains, and its functionality was never verified. Here, we used a reconstitution system in IL-17RA-null fibroblasts to map functional domains within IL-17RA. We demonstrate that the SEFIR domain mediates IL-17RA signaling independently of classic TIR adaptors, such as MyD88 and TRIF. Moreover, we identified a previously undescribed“TIR-like loop” (TILL) required for activation of NF-κB, MAPK, and up-regulation of C/EBPβ and C/EBPδ. Mutagenesis of the TILL domain revealed a site analogous to the LPS d mutation in TLR4, which renders mice insensitive to LPS. However, a putative salt bridge typically found in TIR domains appears to be dispensable. We further identified a C-terminal domain required for activation of C/EBPβ and induction of a subset IL-17 target genes. This structure-function analysis of a IL-17 superfamily receptor reveals important differences in IL-17RA compared with IL-1/TLR receptors.

T lymphocyte (T cell) activation and proliferation is induced by the activation of multiple signal transduction pathways. Earlier studies indicate that CARMA1, a Caspase Recruitment Domain (CARD) and Membrane-associated GUanylate Kinase domain (MAGUK)-containing scaffold protein, plays an essential role in NF-kappaB activation induced by the costimulation of T cell receptor (TCR) and CD28 molecules. However, the molecular mechanism by which CARMA1 mediates TCR-CD28 costimulation-induced NF-kappaB activation is not fully understood. Here we show that CARMA1 is constitutively oligomerized. This oligomerization of CARMA1 is through its Coiled-coil domain. Disruption of the predicted structure of the Coiled-coil domain of CARMA1 impaired its oligomerization and, importantly, abrogated CARMA1-mediated NF-kappaB activation. Interestingly, disruption of the CC1 domain abrogates CARMA1 localization, whereas disruption of the CC2 domain seems to inhibit CARMA1 self-association. Together, our results demonstrate that the oligomerization of CARMA1 is required for TCR-induced NF-kappaB activation.

IL-17 and its receptor are founding members of a novel family of inflammatory cytokines. IL-17 plays a pathogenic role in rheumatoid arthritis (RA)–associated bone destruction. However, IL-17 is also an important regulator of host defense through granulopoiesis and neutrophil trafficking. Therefore, the role of IL-17 in pathogen-initiated bone loss was not obvious. The most common form of infection-induced bone destruction occurs in periodontal disease (PD). In addition to causing significant morbidity, PD is a risk factor for atherosclerotic heart disease and chronic obstructive pulmonary disease (COPD). Similar to RA, bone destruction in PD is caused by the immune response. However, neutrophils provide critical antimicrobial defense against periodontal organisms. Since IL-17 is bone destructive in RA but a key regulator of neutrophils, we examined its role in inflammatory bone loss induced by the oral pathogen Porphyromonas gingivalis in IL-17RA–deficient mice. These mice showed enhanced periodontal bone destruction, suggesting a bone-protective role for IL-17, reminiscent of a neutrophil deficiency. Although IL-17RA–deficient neutrophils functioned normally ex vivo, IL-17RA knock-out (IL-17RAKO) mice exhibited reduced serum chemokine levels and concomitantly reduced neutrophil migration to bone. Consistently, CXCR2KO mice were highly susceptible to alveolar bone loss; interestingly, these mice also suggested a role for chemokines in maintaining normal bone homeostasis. These results indicate a nonredundant role for IL-17 in mediating host defense via neutrophil mobilization.

Sarah Gaffen describes a 2006 paper by Toy et al. that elucidated the subunit composition of the receptor for IL-17.

Disseminated candidiasis is a life threatening nosocomial infection with 40–60% mortality. Fatality from candidiasis is due to fungal invasion of the kidney leading to renal failure. IL-17 is a key cytokine important in the defense against candidiasis, but mechanisms of IL-17-mediated renal immunity remain unexplored. We studied the mechanism of renal protection offered by IL-17 in an intravenous infection model with Candida albicans . We identified a previously unknown role for IL-17 in inducing the renal-protective kallikrein 1(Klk1), a member of a family of serine proteases that constitute the Kallikrein-Kinin system (KKS). IL-17 driven Klk1 is expressed in the kidneys of C. albicans infected kidney. Overexpression of Klk1 and treatment with a bradykinin receptor agonist rescued IL-17RA−/− mice from disseminated candidiasis. Therapeutic manipulation of IL17-KKS pathways using bradykinin prevented apoptosis of renal cells. Moreover, this treatment prevented kidney damage and prolonged survival following C. albicans infection. These data demonstrate that IL-17 driven kallikrein expression contributes to the renal protection against disseminated candidiasis. Thus, exploiting IL-17-KKS pathways with approved drugs may offer avenues for treatment of this fatal infection.

The Interleukin-17A (IL-17A) is related to the development of autoimmune diseases as well as to the control of Candida albicans infections. IL-17 triggers signaling by binding to the IL-17 receptor (IL-17R). The IL-17R recruits adaptor proteins activating downstream effector pathways such as NF-κB and MAPK to induce inflammatory gene expression. To identify IL-17 signaling intermediates we screened candidate genes by siRNA knockdown discovering several novel inhibitors. Based on evidence from human genetic studies and the known importance of A20 as an inhibitor of IL-17R signaling we focused on ABIN1, an A20-interacting protein, which has not previously implicated in this pathway. Since many inhibitors work in a feedback manner to block the pathways that induce their expression, we asked whether IL-17R signaling regulates expression of ABIN1. Indeed, mRNA expression of ABIN1 was induced by IL-17A signaling. Upon ABIN1 knockdown, the IL-17-dependent gene Lipocalin2 (Lcn2) showed increased expression, confirming that ABIN1 is an inhibitor of the IL-17R signaling. Consistently, reconstitution of ABIN1-deficient fibroblast with ABIN1 suppressed Lcn2 expression, and IL-17-induced activation of Lcn2 promoter was downregulated when ABIN1 was ectopically expressed in ST2 cells. However, not all IL-17-dependent target genes were regulated by ABIN1. We are currently dissecting the mechanisms by which ABIN1 inhibits IL-17R signaling. Together, these findings suggest an unexpected complexity in the regulation of IL-17-mediated inflammation.

IL-17A (IL-17) is a proinflammatory cytokine that mediates host defense against extracellular pathogens but also contributes to the pathogenesis of various autoimmune diseases. Despite advances in our understanding of IL-17 signal transduction mechanisms, it is far less clear how IL-17R activity is kept in check to prevent harmful bystander inflammation. In an effort to identify novel, physiologically relevant regulators of IL-17 signal transduction, we evaluated microarray data of IL-17RA-dependent genes that mediate antifungal immunity. We thus identified the gene Zc3h12a (encoding MCPIP1/Regnase-1) as a possible feedback restriction element. MCPIP1 is an endoribonuclease (RNase) that inhibits TLR signaling in part by degrading cytokine mRNA transcripts such as Il6 . Since IL-17 and TLR pathways share multiple target genes including Il6 , we investigated the role of MCPIP1 in IL-17 signal transduction. Here, we demonstrate that MCPIP1 is upregulated by IL-17 and serves as a potent feedback inhibitor of IL-17 signal transduction. IL-17 signaling inhibition required MCPIP1’s RNase activity, but not its deubiquitinase activity. In vivo , resistance to disseminated Candida albicans infection was enhanced in MCPIP1 haploinsufficient mice; resistance was reversed in an IL-17R-deficient background. Conversely, IL-17-dependent pathology in MCPIP1+/− mice was dramatically exacerbated during EAE and pulmonary inflammation. In vitro , MCPIP1 strongly inhibited the Lcn2 promoter activation by regulating the mRNA stability of the I κ B ζ transcription factor. Unexpectedly, MCPIP1 also degraded Il17ra and Il17rc mRNA, independently of the 3’ UTR. The cumulative impact of MCPIP1 on IL-6, I κ B ζ and possibly IL-17R subunits results in a biologically relevant inhibition of IL-17 signaling.

Humans or mice subjected to immunosuppression, such as corticosteroids or anti-cytokine biologic therapies, are susceptible to mucosal infections by the commensal fungus Candida albicans. Recently it has become evident that the Th17/IL-17 axis is essential for immunity to candidiasis, but the downstream events that control immunity to this fungus are poorly understood. The CCAAT/Enhancer Binding Protein-β (C/EBPβ) transcription factor is important for signaling by multiple inflammatory stimuli, including IL-17. C/EBPβ is regulated in a variety of ways by IL-17, and controls several downstream IL-17 target genes. However, the role of C/EBPβ in vivo is poorly understood, in part because C/EBPβ-deficient mice are challenging to breed and work with. In this study, we sought to understand the role of C/EBPβ in the context of an IL-17-dependent immune response, using C. albicans infection as a model system. Confirming prior findings, we found that C/EBPβ is required for immunity to systemic candidiasis. In contrast, C/EBPβ(-/-) mice were resistant to oropharyngeal candidiasis (OPC), in a manner indistinguishable from immunocompetent WT mice. However, C/EBPβ(-/-) mice experienced more severe OPC than WT mice in the context of cortisone-induced immunosuppression. Expression of the antimicrobial peptide β-defensin (BD)-3 correlated strongly with susceptibility in C/EBPβ(-/-) mice, but no other IL-17-dependent genes were associated with susceptibility. Therefore, C/EBPβ contributes to immunity to mucosal candidiasis during cortisone immunosuppression in a manner linked to β-defensin 3 expression, but is apparently dispensable for the IL-17-dependent response.

Huppler, A. R., H. R. Conti, N. Hernandez-Santos, T. Darville, P. S. Biswas, and S. L. Gaffen. 2014. Role of neutrophils in IL-17–dependent immunity to mucosal candidiasis. J. Immunol. 192: [1745–1752][1]. There was an inadvertent error in the calculation of the absolute neutrophil count due