T helper cells that produce interleukin 17 (IL-17) are associated with inflammation and the control of certain bacteria. We report here the essential involvement of the adaptor protein Act1 in IL-17 receptor (IL-17R) signaling and IL-17-dependent immune responses. After stimulation with IL-17, recruitment of Act1 to IL-17R required the IL-17R conserved cytoplasmic 'SEFIR' domain, followed by recruitment of the kinase TAK1 and E3 ubiquitin ligase TRAF6, which mediate 'downstream' activation of transcription factor NF-kappaB. IL-17-induced expression of inflammation-related genes was abolished in Act1-deficient primary astroglial and gut epithelial cells. This reduction was associated with much less inflammatory disease in vivo in both autoimmune encephalomyelitis and dextran sodium sulfate-induced colitis. Our data show that Act1 is essential in IL-17-dependent signaling in autoimmune and inflammatory disease.

SUMMARY Interleukin-17 (IL-17) secreted by T helper 17 (Th17) cells is essential in the development of experimental autoimmune encephalomyelitis (EAE). However, it remains unclear how IL-17-mediated signaling in different cellular compartments participates in the central nervous system (CNS) inflammatory process. We examined CNS inflammation in mice with specific deletion of Act1, a critical component required for IL-17 signaling, in endothelial cells, macrophages and microglia, and neuroectoderm (neurons, astrocytes, and oligodendrocytes). In Act1-deficient mice, Th17 cells showed normal infiltration into the CNS but failed to recruit lymphocytes, neutrophils, and macrophages. Act1 deficiency in endothelial cells or in macrophages and microglia did not substantially impact the development of EAE. However, targeted Act1 deficiency in neuroectoderm derived CNS resident cells resulted in markedly reduced severity in EAE. Specifically, Act1-deficient astrocytes showed impaired IL-17-mediated inflammatory gene induction. Thus, astroctyes are critical in IL-17-Act1 mediated leukocyte recruitment during autoimmune induced inflammation of the CNS.

Interleukin (IL)-17, a proinflammatory cytokine mainly produced by T-helper-17 (T H 17) lineage, is required for host defense against bacteria and fungus infection and plays a critical role in the pathogenesis of inflammatory and autoimmune diseases. Act1 is an essential adaptor molecule in IL-17-mediated signaling pathway, recruited to IL-17 receptor (IL-17R) upon IL-17 stimulation through SEFIR-SEFIR domain interaction. Here we report that Act1 is a novel bona fide U-box E3 ubiquitin ligase, whose activity is essential for IL-17-mediated signaling pathways (including nuclear factor kappa B (NFκB), and partially required for Jun N-terminal Kinase (JNK) and extracellular signal-regulated kinase (ERK) activation) and inflammatory gene expression (KC (CXCL1), granulocyte macrophage colony stimulating factor (GM-CFS ) and IL-6) in mammalian cells. By utilizing Ubc13/Uev1A E2 complex, Act1 mediates Lys 63-linked ubiquitination of tumor necrosis factor receptor-associated factor 6 (TRAF6), an important signaling component of IL-17-mediated signaling pathway. Deletion and point mutations of the Act1 U-box abolish Act1-mediated ubiquitination of TRAF6 and impair the ability of Act1 to restore IL-17-dependent signaling and inflammatory gene expression in Act1 −/− mouse embryonic fibroblasts (MEFs). Importantly, we demonstrate that the Lys 124 residue of TRAF6 is critical for efficient Act1-mediated TRAF6 ubiquitination and for the ability of TRAF6 to mediate IL-17-induced NFκB activation. Thus Act1 mediates IL-17-induced signaling pathways through its E3 ubiquitin ligase activity and TRAF6 is a critical substrate of Act1, indicating the importance of protein ubiquitination in IL-17-dependent inflammatory response.

DCIR2+ DCs can initiate extrafollicular B cell responses to T cell–dependent antigen.

Transgenic expression of TLR7 results in the expansion and hyperactivation of T1 B cells in response to endogenous RNA complexes, leading to increased autoantibody production.

Neutral sphingomyelinase (NSMase) has been proposed to mediate interleukin (IL)-1beta signaling in liver. In this paper, we used adenovirus-mediated gene transfer to inducibly express FLAG-tagged mouse NSMase-2 in primary rat hepatocytes in order to further elucidate the molecular nature of the NSMase involved. Initial studies confirmed that the EST clone used in these experiments encoded a Mg2+-dependent NSMase. The in vitro activity of the heterologously expressed enzyme was inhibited in the presence of 0.5% Triton or 50 mM EDTA. In addition, the expression of this NSMase-2 clone in primary hepatocytes led to increased cellular levels of ceramide, indicating that the enzyme is active in situ. Immunofluorescence studies in Hep G2 cells infected with NSMase-2 expressing adenoviruses showed that the FLAG-tagged NSMase-2 was localized to the plasma membrane. Cell viability remained unchanged 72 h following infection and induction. The effect of NSMase-2 expression on IL-1beta-induced activation of c-Jun N-terminal kinase (JNK) was tested. Expression of NSMase-2 increased JNK phosphorylation between 1.5- and 2-fold over the basal level. Furthermore, NSMase-2 was found to strongly increase the ability of IL-1beta to phosphorylate JNK. This potentiation was mediated by a phosphatase from the PP2A family, possibly by modulating the phosphorylation pattern of IL-1beta receptor-associated kinase (IRAK). In conclusion, the data presented suggest that NSMase-2 could be involved in IL-1beta-induced JNK activation in hepatocytes.

Acid sphingomyelinase plays important roles in ceramide homeostasis, which has been proposed to be linked to insulin resistance. To test this association in vivo, acid sphingomyelinase deletion (asm ؊/؊ ) was transferred to mice lacking the low density lipoprotein receptor (ldlr ؊/؊ ), and then offsprings were placed on control or modified (enriched in saturated fat and cholesterol) diets for 10 weeks. The modified diet caused hypercholesterolemia in all genotypes; however, in contrast to asm ؉/؉ /ldlr ؊/؊ , the acid sphingomyelinase-deficient littermates did not display hepatic triacylglyceride accumulation, although sphingomyelin and other sphingolipids were substantially elevated, and the liver was enlarged. asm ؊/؊ /ldlr ؊/؊ mice on a modified diet did not accumulate body fat and were protected against diet-induced hyperglycemia and insulin resistance. Experiments with hepatocytes revealed that acid sphingomyelinase regulates the partitioning of the major fatty acid in the modified diet, palmitate, into two competitive and inversely related pools, triacylglycerides and sphingolipids, apparently via modulation of serine palmitoyltransferase, a rate-limiting enzyme in de novo sphingolipid synthesis. These studies provide evidence that acid sphingomyelinase activity plays an essential role in the regulation of glucose metabolism by regulating the hepatic accumulation of triacylglycerides and sphingolipids during consumption of a diet rich in saturated fats.

CD22 (Siglec 2) is a receptor predominantly restricted to B cells. It was initially characterized over 30 years ago and named “CD22” in 1984 at the 2nd International workshop in Boston (1). Several excellent reviews have detailed CD22 functions, CD22-regulated signaling pathways and B cell subsets regulated by CD22 or Siglec G (2–4). This review is an attempt to highlight recent and possibly forgotten findings. We also describe the role of CD22 in autoimmunity and the great potential for CD22-based immunotherapeutics for the treatment of autoimmune diseases such as systemic lupus erythematosus (SLE).