MicroRNA-142 Is Critical for the Homeostasis and Function of Type 1 Innate Lymphoid Cells.

Natural killer (NK) cells are cytotoxic type 1 innate lymphoid cells (ILCs) that defend against viruses and mediate anti-tumor responses, yet mechanisms controlling their development and function remain incompletely understood. We hypothesized that the abundantly expressed microRNA-142 (miR-142) is a critical regulator of type 1 ILC biology. Interleukin-15 (IL-15) signaling induced miR-142 expression, whereas global and ILC-specific miR-142-deficient mice exhibited a cell-intrinsic loss of NK cells. Death of NK cells resulted from diminished IL-15 receptor signaling within miR-142-deficient mice, likely via reduced suppressor of cytokine signaling-1 (Socs1) regulation by miR-142-5p. ILCs persisting in Mir142-/- mice demonstrated increased expression of the miR-142-3p target αV integrin, which supported their survival. Global miR-142-deficient mice exhibited an expansion of ILC1-like cells concurrent with increased transforming growth factor-ß (TGF-ß) signaling. Further, miR-142-deficient mice had reduced NK-cell-dependent function and increased susceptibility to murine cytomegalovirus (MCMV) infection. Thus, miR-142 critically integrates environmental cues for proper type 1 ILC homeostasis and defense against viral infection.

The signaling suppressor CIS controls proallergic T cell development and allergic airway inflammation.

Transcription factors of the STAT family are critical in the cytokine-mediated functional differentiation of CD4(+) helper T cells. Signaling inhibitors of the SOCS family negatively regulate the activation of STAT proteins; however, their roles in the differentiation and function of helper T cells are not well understood. Here we found that the SOCS protein CIS, which was substantially induced by interleukin 4 (IL-4), negatively regulated the activation of STAT3, STAT5 and STAT6 in T cells. CIS-deficient mice spontaneously developed airway inflammation, and CIS deficiency in T cells led to greater susceptibility to experimental allergic asthma. CIS-deficient T cells showed enhanced differentiation into the TH2 and TH9 subsets of helper T cells. STAT5 and STAT6 regulated IL-9 expression by directly binding to the Il9 promoter. Our data thus demonstrate a critical role for CIS in controlling the proallergic generation of helper T cells.

The microRNA miR-155 controls CD8(+) T cell responses by regulating interferon signaling.

We found upregulation of expression of the microRNA miR-155 in primary effector and effector memory CD8(+) T cells, but low miR-155 expression in naive and central memory cells. Antiviral CD8(+) T cell responses and viral clearance were impaired in miR-155-deficient mice, and this defect was intrinsic to CD8(+) T cells, as miR-155-deficient CD8(+) T cells mounted greatly diminished primary and memory responses. Conversely, miR-155 overexpression augmented antiviral CD8(+) T cell responses in vivo. Gene-expression profiling showed that miR-155-deficient CD8(+) T cells had enhanced type I interferon signaling and were more susceptible to interferon's antiproliferative effect. Inhibition of the type I interferon-associated transcription factors STAT1 or IRF7 resulted in enhanced responses of miR-155-deficient CD8(+) T cells in vivo. We have thus identified a previously unknown role for miR-155 in regulating responsiveness to interferon and CD8(+) T cell responses to pathogens in vivo.

Ubc13 maintains the suppressive function of regulatory T cells and prevents their conversion into effector-like T cells.

The maintenance of immune homeostasis requires regulatory T cells (Treg cells). Here we found that Treg cell­specific ablation of Ubc13, a Lys63 (K63)-specific ubiquitin-conjugating enzyme, caused aberrant T cell activation and autoimmunity. Although Ubc13 deficiency did not affect the survival of Treg cells or expression of the transcription factor Foxp3, it impaired the in vivo suppressive function of Treg cells and rendered them sensitive to the acquisition of T helper type 1 (TH1) cell­ and interleukin 17 (IL-17)-producing helper T (TH17) cell­like effector phenotypes. This function of Ubc13 involved its downstream target, the kinase IKK. The Ubc13-IKK signaling axis controlled the expression of specific Treg cell effector molecules, including IL-10 and SOCS1. Collectively, our findings suggest that the Ubc13-IKK signaling axis regulates the molecular program that maintains Treg cell function and prevents Treg cells from acquiring inflammatory phenotypes.

Genome-wide identification and characterization of large yellow croaker (Larimichthys crocea) suppressors of cytokine signaling (SOCS) in immune response to Pseudomonas plecoglossicida infection and acute hypoxia stress.

The suppressor of cytokine signaling (SOCS) gene family is a group of genes involved in the negative regulation of cytokine signal transduction. The members of this family play a crucial role in regulating immune and inflammatory processes. However, comprehensive investigations of these genes have not yet been conducted in the economically significant fish large yellow croaker (Larimichthys crocea). In this study, a total of 13 SOCS genes (LcSOCS1a, LcSOCS1b, LcSOCS2, LcSOCS3a, LcSOCS3b, LcSOCS4, LcSOCS5a, LcSOCS5b, LcSOCS6, LcSOCS7a, LcSOCS7b, LcCISHa and LcCISHb) were identified and analyzed in L. crocea. The phylogenetic tree revealed a high conservation of SOCS genes in evolution, and the gene structure and motif analysis indicated a high similarity in the structure of LcSOCSs in the same subfamily. In addition, the expression patterns of LcSOCSs showed that LcSOCS1b was significantly down-regulated in all time under acute hypoxia stress, but it was markedly up-regulated throughout the entire process after P. plecoglossicida infection, revealing its different immune effects to two stresses. Besides, LcSOCS2a, LcSOCS6 and LcSOCS7a only participated in acute hypoxic stress, while LcSOCS5a was more sensitive to P. plecoglossicida infection. In summary, these results indicated that SOCS genes were involved in stress responses to both biological and non-biological stimuli, setting the foundation for deeper study on the functions of SOCS genes.

An unconventional role of an ASB family protein in NF-κB activation and inflammatory response during microbial infection and colitis.

Nuclear factor κB (NF-κB)-mediated signaling pathway plays a crucial role in the regulation of inflammatory process, innate and adaptive immune responses. The hyperactivation of inflammatory response causes host cell death, tissue damage, and autoinflammatory disorders, such as sepsis and inflammatory bowel disease. However, how these processes are precisely controlled is still poorly understood. In this study, we demonstrated that ankyrin repeat and suppressor of cytokine signaling box containing 1 (ASB1) is involved in the positive regulation of inflammatory responses by enhancing the stability of TAB2 and its downstream signaling pathways, including NF-κB and mitogen-activated protein kinase pathways. Mechanistically, unlike other members of the ASB family that induce ubiquitination-mediated degradation of their target proteins, ASB1 associates with TAB2 to inhibit K48-linked polyubiquitination and thereby promote the stability of TAB2 upon stimulation of cytokines and lipopolysaccharide (LPS), which indicates that ASB1 plays a noncanonical role to further stabilize the target protein rather than induce its degradation. The deficiency of Asb1 protects mice from Salmonella typhimurium- or LPS-induced septic shock and increases the survival of mice. Moreover, Asb1-deficient mice exhibited less severe colitis and intestinal inflammation induced by dextran sodium sulfate. Given the crucial role of ASB proteins in inflammatory signaling pathways, our study offers insights into the immune regulation in pathogen infection and inflammatory disorders with therapeutic implications.

Role of Suppressor of cytokine signaling 2 during the development and resolution of an experimental arthritis.

Rheumatoid arthritis(RA) is a debilitating chronic inflammatory disease. Suppressors of Cytokine Signaling(SOCS) proteins regulate homeostasis and pathogenesis in several diseases. The intersection between RA pathophysiology and SOCS2 is unclear. Herein, we investigated the roles of SOCS2 during the development of an experimental antigen-induced arthritis(AIA). In wild type mice, joint SOCS2 expression was reduced during AIA development. At the peak of inflammation, SOCS2-/- mice presented with reduced numbers of infiltrated cells in their joints. At the late phase of AIA, however, exhibited increased adhesion/infiltration of neutrophils, macrophages, CD4+-T cells, CD4+CD8+-T cells, and CD4-CD8--T cells associated with elevated IL-17 and IFN-γ levels, joint damage, proteoglycan loss, and nociception. SOCS2 deficiency resulted in lower numbers of apoptotic neutrophils and reduced efferocytosis. The present study demonstrated the vital role of SOCS2 during the development and resolution of an experimental RA model. Hence, this protein may be a novel therapeutic target for this disorder.

SOCS: negative regulators of cytokine signaling for immune tolerance.

Cytokines are important intercellular communication tools for immunity. Many cytokines promote gene transcription and proliferation through the JAK/STAT (Janus kinase/signal transducers and activators of transcription) and the Ras/ERK (GDP/GTP-binding rat sarcoma protein/extracellular signal-regulated kinase) pathways, and these signaling pathways are tightly regulated. The SOCS (suppressor of cytokine signaling) family members are representative negative regulators of JAK/STAT-mediated cytokine signaling and regulate the differentiation and function of T cells, thus being involved in immune tolerance. Human genetic analysis has shown that SOCS family members are strongly associated with autoimmune diseases, allergy and tumorigenesis. SOCS family proteins also function as immune-checkpoint molecules that contribute to the unresponsiveness of T cells to cytokines.

Suppressors of cytokine signaling 2 and 3 diametrically control macrophage polarization.

Suppressors of cytokine signaling (SOCS) are important regulators of lipopolysaccharide (LPS) and cytokine responses but their role in macrophage polarization is unknown. We have shown here that myeloid-restricted Socs3 deletion (Socs3(Lyz2cre)) resulted in resistance to LPS-induced endotoxic shock, whereas Socs2(-/-) mice were highly susceptible. We observed striking bias toward M2-like macrophages in Socs3(Lyz2cre) mice, whereas the M1-like population was enriched in Socs2(-/-) mice. Adoptive transfer experiments showed that responses to endotoxic shock and polymicrobial sepsis were transferable and macrophage dependent. Critically, this dichotomous response was associated with enhanced regulatory T (Treg) cell recruitment by Socs3(Lyz2cre) cells, whereas Treg cell recruitment was absent in the presence of Socs2(-/-) macrophages. In addition, altered polarization coincided with enhanced interferon-gamma (IFN-γ)-induced signal transducer and activator of transcription-1 (STAT1) activation in Socs2(-/-) macrophages and enhanced interleukin-4 (IL-4) plus IL-13-induced STAT6 phosphorylation in Socs3(Lyz2cre) macrophages. SOCS, therefore, are essential controllers of macrophage polarization, regulating inflammatory responses.

An interleukin-21-interleukin-10-STAT3 pathway is critical for functional maturation of memory CD8+ T cells.

Memory CD8(+) T cells are critical for long-term immunity, but the genetic pathways governing their formation remain poorly defined. This study shows that the IL-10-IL-21-STAT3 pathway is critical for memory CD8(+) T cell development after acute LCMV infection. In the absence of either interleukin-10 (IL-10) and IL-21 or STAT3, virus-specific CD8(+) T cells retain terminal effector (TE) differentiation states and fail to mature into protective memory T cells that contain self-renewing central memory T cells. Expression of Eomes, BCL-6, Blimp-1, and SOCS3 was considerably reduced in STAT3-deficient memory CD8(+) T cells, and BCL-6- or SOCS3-deficient CD8(+) T cells also had perturbed memory cell development. Reduced SOCS3 expression rendered STAT3-deficient CD8(+) T cells hyperresponsive to IL-12, suggesting that the STAT3-SOCS3 pathway helps to insulate memory precursor cells from inflammatory cytokines that drive TE differentiation. Thus, memory CD8(+) T cell precursor maturation is an active process dependent on IL-10-IL-21-STAT3 signaling.

Leukemia inhibitory factor inhibits T helper 17 cell differentiation and confers treatment effects of neural progenitor cell therapy in autoimmune disease.

Neural progenitor cell (NPC) therapy is considered a promising treatment modality for multiple sclerosis (MS), potentially acting through neural repair. Here, we showed that intravenous administration of NPCs ameliorated experimental autoimmune encephalomyelitis (EAE) by selectively inhibiting pathogenic T helper 17 (Th17) cell differentiation. Leukemia inhibitory factor (LIF) produced by NPCs was responsible for the observed EAE suppression. Through the inducible LIF receptor expression, LIF inhibited the differentiation of Th17 cells in EAE mice and that from MS subjects. At the molecular level, LIF exerted an opposing effect on interleukin 6 (IL-6)-induced signal transducer and activator of transcription 3 (STAT3) phosphorylation required for Th17 cell differentiation by triggering a signaling cascade that activated extracellular signal-regulated MAP kinase (ERK) and upregulated suppressor of cytokine signaling 3 (SOCS3) expression. This study reveals a critical role for LIF in regulating Th17 cell differentiation and provides insights into the mechanisms of action of NPC therapy in MS.

Molecular characterization, expression pattern and evolution of nine suppressors of cytokine signaling (SOCS) gene in the swamp eel (Monopterus albus).

Suppressors of cytokine signaling (SOCS) family members have negative effects on cytokine signaling pathways involved in immunity, growth and development. Owing to their typical feature, they have been extensively studied in mammalians, but they have not offered systematic studies among teleosts. In the present study, nine SOCS family genes were identified in the swamp eel genome and analyzed regulation mechanisms of SOCS family members in swamp eels. The open reading frames of MaSOCS1a, MaSOCS1b, MaSOCS2, MaSOCS3a, MaSOCS3b, MaSOCS4, MaSOCS5, MaSOCS6 and MaSOCS7 were 663 bp, 603 bp, 717 bp, 618 bp, 645 bp, 1188 bp, 1488 bp, 1611 bp and 1998 bp and encoded 220, 238, 200, 205, 214, 395, 496, 536 and 655 amino acids, respectively. All SOCS proteins have no signal peptides. Multiple alignment revealed that MaSOCS family members possessed a typical conserved SOCS box and SH2 region. Phylogenetic analyses showed that all SOCS proteins were divided into two main clusters. Taken together with the similarity and identity of SOCS protein amino acids, these results indicated that MaSOCS family members shared conserved with other homologous genes, in which MaSOCS7 was more conserved. Further syntenic analysis confirmed the phylogenetic analysis results and annotation of SOCS protein, suggesting that MaSOCS5 shared a common ancestor gene with that of fish and humans. MaSOCS family members were constitutively expressed in a wide range of tissues with different levels. In particular, spleen and head kidneys play an important role in immune-related pathways. After Aeromonas veronii and polyinosinic-polycytidylic acid (poly I:C) challenge in the spleen and head kidney, MaSOCS family members exhibit different expression profiles. These expression patterns indicated that MaSOCS family members could make acute responses after pathogen invasion. Taken together, these results indicate that MaSOCS family members participate in the immune response against pathogens and offer a solid foundation for future studies of SOCS function.

SOCS-mediated immunomodulation of natural killer cells.

Natural killer (NK) cells are innate immune cells with an intrinsic ability to detect and kill infected and cancerous cells. The success of therapies targeting immune checkpoints on CD8 cells has intensified interest in harnessing the cytolytic effector functions of NK cells for new cancer treatments. NK cell development, survival and effector activity is dependent on exposure to the cytokine interleukin (IL)-15. The suppressor of cytokine (SOCS) proteins (CIS; SOCS1-7) are important negative regulators of cytokine signaling, and both CIS and SOCS2 are reported to have roles in regulating NK cell responses. Their immunomodulatory effects on NK cells suggest that these SOCS proteins are promising targets that can potentially form the basis of novel cancer therapies. Here we discuss the role of NK cells in tumor immunity as well as review the role of the SOCS proteins in regulating IL-15 signaling and NK cell function.

Molecular characterization of nine suppressors of cytokine signaling (SOCS) genes from yellow catfish Pelteobagrus fulvidraco and their changes in mRNA expression to dietary carbohydrate levels.

Suppressors of cytokine signaling (SOCS) are important molecules that mediates the regulation of glucose homeostasis. Here, we cloned and characterized the full-length cDNA sequences of nine genes of the SOCS family (SOCS1, 2, 3, 3b, 5, 5b, 6, 7 and CISH) from yellow catfish P. fulvidraco, explored their mRNA abundance across the tissues and their mRNA changes to dietary carbohydrate levels. Structural analysis indicated that the nine members shared conserved functional domains to the orthologues of the mammalian SOCS members, such as SRC homology 2 and the SOCS domains. Their mRNAs were constitutively expressed in various tissues but changed among the tissues. Their mRNA expression in response to dietary carbohydrate levels were explored in the liver, muscle, intestine, testis and ovary. Dietary carbohydrate addition showed significant effects on the mRNA levels of the nine SOCS members. Moreover, their mRNA expressions in response to dietary carbohydrate levels were also tissue-dependent. These indicated that SOCS members potentially mediated the utilization of dietary carbohydrate in yellow catfish.

CgSOCS6 negatively regulates the expression of CgIL17s and CgDefh1 in the pacific oyster Crassostrea gigas.

As a family of negatively feedback regulating factors, the suppressor of cytokine signaling (SOCS) can depress cytokine signal transduction, and eventually modulate growth, development, differentiation, and immune response. In the present study, a SOCS homologue (designated as CgSOCS6) was identified from oyster Crassostrea gigas. The open reading frame of CgSOCS6 cDNA was of 1167 bp encoding a peptide of 388 amino acid residues with a central Src homology 2 (SH2) domain, a conserved C-terminal SOCS box, and a nucleus localization sequence (NLS) in its N-terminus. The deduced amino acid sequence of CgSOCS6 shared 37.9-45.5% similarity with other SOCS6/7 family members. In the unrooted phylogenetic tree, CgSOCS6 was clustered with EsSOCS6 from Chinese mitten crab Eriocheir sinensis and assigned into the SOCS6/7 group. The mRNA transcripts of CgSOCS6 were constitutively distributed in all the tested tissues, with the highest level in hemocytes. After lipopolysaccharide (LPS) stimulation, the mRNA expression of CgSOCS6 in hemocytes was significantly up-regulated to the highest level at 6 h (8.48-fold compared to the control group, p < 0.01), and then kept at a relatively higher level from 12 h to 72 h. CgSOCS6 protein could be translocated into the hemocyte nucleus after LPS stimulation. The mRNA expressions of interleukin 17-4 (CgIL17-4), CgIL17-5, and defensin (CgDefh1) in the hemocytes of CgSOCS6-knockdown oysters increased significantly (2.55-fold, 2.68-fold, 4.68-fold of that in EGFP-RNAi oysters, p < 0.05, p < 0.05, p < 0.001, respectively) after LPS stimulation. These findings suggested that CgSOCS6 was involved in the oyster immune response by regulating the expressions of CgIL17-4, CgIL17-5, and CgDefh1.

TLR7 Signaling Regulates Th17 Cells and Autoimmunity: Novel Potential for Autoimmune Therapy.

Innate regulation through TLR signaling has been shown to be important for promoting T cell subset development and function. However, limited information is known about whether differential TLR signaling can selectively inhibit Th17 and/or Th1 cells, which are important for controlling excessive inflammation and autoimmune responses. In this article, we demonstrate that activation of TLR7 signaling in T cells can inhibit Th17 cell differentiation from naive T cells and IL-17 production in established Th17 cells. We further report that downregulation of STAT3 signaling is responsible for TLR7-mediated inhibition of Th17 cells due to induction of suppressor of cytokine signaling 3 and 5. TLR7-mediated suppression of Th17 cells does not require dendritic cell involvement. In addition, we show that TLR7 signaling can suppress Th1 cell development and function through a mechanism different from Th17 cell suppression. Importantly, our complementary in vivo studies demonstrate that treatment with the TLR7 ligand imiquimod can inhibit Th1 and Th17 cells, resulting in the prevention of, and an immunotherapeutic reduction in, experimental autoimmune encephalomyelitis. These studies identify a new strategy to manipulate Th17/Th1 cells through TLR7 signaling, with important implications for successful immunotherapy against autoimmune and inflammatory diseases.

The kinase Akt1 controls macrophage response to lipopolysaccharide by regulating microRNAs.

MicroRNAs regulated by lipopolysaccharide (LPS) target genes that contribute to the inflammatory phenotype. Here, we showed that the protein kinase Akt1, which is activated by LPS, positively regulated miRNAs let-7e and miR-181c but negatively regulated miR-155 and miR-125b. In silico analyses and transfection studies revealed that let-7e repressed Toll-like receptor 4 (TLR4), whereas miR-155 repressed SOCS1, two proteins critical for LPS-driven TLR signaling, which regulate endotoxin sensitivity and tolerance. As a result, Akt1(-/-) macrophages exhibited increased responsiveness to LPS in culture and Akt1(-/-) mice did not develop endotoxin tolerance in vivo. Overexpression of let-7e and suppression of miR-155 in Akt1(-/-) macrophages restored sensitivity and tolerance to LPS in culture and in animals. These results indicate that Akt1 regulates the response of macrophages to LPS by controlling miRNA expression.

The involvement of suppressor of cytokine signaling 6 (SOCS6) in immune response of Chinese mitten crab Eriocheir sinensis.

Suppressor of cytokine signaling (SOCS) is a family of cytokine-inducible negative regulators of cytokine signaling and it plays a crucial role in various physiological processes. In the present study, the full-length cDNA of a SOCS (designated as EsSOCS6) was cloned from Chinese mitten crab Eriocheir sinensis. The open reading frame of EsSOCS6 cDNA was of 1266 bp, which encoded a polypeptide of 421 amino acid residues. There were two typically conserved SOCS family domains in EsSOCS6, including a central Src homology 2 (SH2) domain and a C-terminal SOCS box. The deduced amino acid sequence of EsSOCS6 shared 72-76% similarity with those of other SOCS6 family members. EsSOCS6 mRNA was constitutively expressed in all the examined tissues with higher expression levels in the immune-related tissues, such as hepatopancreas, hemocytes and gill. The mRNA expression levels of the EsSOCS6 in hemocytes were significantly up-regulated after the stimulations with lipopolysaccharide (LPS), Aeromonas hydrophila and polyinosinic-polycytidylic acid (poly (I:C)). The mRNA expressions of threonine/serine protein kinase (EsAkt) and EsRelish were dramatically declined after EsSOCS6 was interfered by dsRNA. Collectively, these results demonstrated that EsSOCS6 might regulate the activation of the NF-κB signaling pathway and play an important role in the innate immune responses of E. sinensis.

Evolution of Cytokine Receptor Signaling.

Cytokines represent essential mediators of cell-cell communication with particularly important roles within the immune system. These secreted factors are produced in response to developmental and/or environmental cues and act via cognate cytokine receptors on target cells, stimulating specific intracellular signaling pathways to facilitate appropriate cellular responses. This review describes the evolution of cytokine receptor signaling, focusing on the class I and class II receptor families and the downstream JAK-STAT pathway along with its key negative regulators. Individual components generated over a long evolutionary time frame coalesced to form an archetypal signaling pathway in bilateria that was expanded extensively during early vertebrate evolution to establish a substantial "core" signaling network, which has subsequently undergone limited diversification within discrete lineages. The evolution of cytokine receptor signaling parallels that of the immune system, particularly the emergence of adaptive immunity, which has likely been a major evolutionary driver.