The IL-6 family of cytokines modulates STAT3 activation by desumoylation of PML through SENP1 induction

Post-translational modification by small ubiquitin-like modifier (SUMO) plays an important role in the regulation of different signaling pathways and is involved in the formation of promyelocytic leukemia (PML) protein nuclear bodies following sumoylation of PML. In the present study, we found that IL-6 induces desumoylation of PML and dissociation between PML and SUMO1 in hepatoma cells. We also found that IL-6 induces mRNA expression of SENP1, a member of the SUMO-specific protease family. Furthermore, wild-type SENP1 but not an inactive SENP1 mutant restored the PML-mediated suppression of STAT3 activation. These results indicate that the IL-6 family of cytokines modulates STAT3 activation by desumoylation and inactivation PML through SENP1 induction.     

Interleukin-6 family of cytokines mediate angiotensin II-induced cardiac hypertrophy in rodent cardiomyocytes

This study was designed to investigate whether angiotensin II induces the interleukin (IL)-6 family of cytokines in cardiac fibroblasts and, if so, whether these cytokines can augment cardiac hypertrophy. Angiotensin II increased IL-6, leukemia inhibitory factor (LIF) and cardiotrophin-1 mRNA by 6.5-, 10.2-, and 2.0-fold, respectively, but did not affect IL-11, ciliary neurotrophic factor, or oncostatin M in cardiac fibroblasts. Enzyme-linked immunosorbent assay revealed that angiotensin II-stimulated conditioned medium from cardiac fibroblasts contained 9.3 ng/ml IL-6 at 24 h, which was 24-fold higher than the control. It phosphorylated gp130 and STAT3 in cardiomyocytes, which was reduced with RX435 (anti-gp130 blocking antibody). It increased [(3)H]phenylalanine uptake and cell area by 44% and 86% in cardiomyocytes compared with mock medium. RX435 suppressed these increases by 26% and 38%, while TAK044 (endothelin-A/B-R blocker) suppressed them by 52% and 52%, respectively. Antisense oligonucleotides against LIF and cardiotrophin-1 blocked their up-regulation, and attenuated the conditioned medium-induced increase in [(3)H]phenylalanine uptake by 21% and 13%, respectively. The combination of antisense oligonucleotides to LIF and cardiotrophin-1 decreased their uptake by 33%. These results indicated that angiotensin II induced IL-6, LIF, and cardiotrophin-1 in cardiac fibroblasts, and that these cytokines, particularly LIF and cardiotrophin-1, activated gp130-linked signaling and contributed to angiotensin II-induced cardiomyocyte hypertrophy.     

Autocrine IL-6/STAT3 signaling aids development of acquired drug resistance in Group 3 medulloblastoma

Medulloblastoma (MB) is a high-grade pediatric brain malignancy that originates from neuronal precursors located in the posterior cranial fossa. In this study, we evaluated the role of STAT3 and IL-6 in a tumor microenvironment mediated drug resistance in human MBs. We established that the Group 3 MB cell line, Med8A, is chemosensitive (hence Med8A-S), and this is correlated with a basal low phosphorylated state of STAT3, while treatment with IL-6 induced robust increases in pY705-STAT3. Via incremental selection with vincristine, we derived the stably chemoresistant variant, Med8A-R, that exhibited multi-drug resistance, enhanced IL-6 induced pY705-STAT3 levels, and increased IL6R expression. Consequently, abrogation of STAT3 or IL6R expression in Med8A-R led to restored chemosensitivity to vincristine, highlighting a prominent role for canonical IL-6/STAT3 signaling in acquired drug resistance. Furthermore, Med8A-S subjected to conditioning exposure with IL-6, termed Med8A-IL6+ cells, exhibited enhanced vincristine resistance, increased expression of pY705-STAT3 and IL6R, and increased secretion of IL-6. When cocultured with Med8A-IL6+ cells, Med8A-S cells exhibited increased pY705-STAT3 and increased IL-6 secretion, suggesting a cytokine feedback loop responsible for amplifying STAT3 activity. Similar IL-6 induced phenomena were also observed in the Group 3 MB cell lines, D283 and D341, including increased pY705-STAT3, drug resistance, IL-6 secretion and IL6R expression. Our study unveiled autocrine IL-6 as a promoter of STAT3 signaling in development of drug resistance, and suggests therapeutic benefits for targeting the IL-6/STAT3 signaling axis in Group 3 MBs.Subject terms: Cancer microenvironment, CNS cancer, Paediatric cancer     

Mechanisms of unconventional secretion of IL-1 family cytokines

One of the most poorly understood processes in cell biology is the peculiar ability of specific leaderless proteins to be secreted via ER/Golgi-independent mechanisms (‘unconventional protein secretion’). One such leaderless protein is the major immune-activating cytokine, interleukin-1β (IL-1β). Unusual amongst cytokines, IL-1β is expressed in the cytosol as an inactive precursor protein. It requires maturation by the caspase-1 protease, which itself requires activation upon immune cell sensing of infection or cell stress. Despite 25 years of intensive research into IL-1β secretory mechanisms, how it exits the cell is still not well understood. Here we will review the various mechanisms by which macrophages have been proposed to secrete IL-1 family cytokines, and the potential involvement of caspase-1 therein. Since aberrant IL-1β production drives inherited and acquired human diseases (e.g. autoinflammatory diseases, arthritic diseases, gout, Alzheimer’s disease), elucidation of the IL-1β secretory pathway may offer new therapeutic opportunities for treatment across this wide range of human conditions.     

IL-6 regulates in vivo dendritic cell differentiation through STAT3 activation

Dendritic cells (DCs) orchestrate immune responses according to their state of maturation. In response to infection, DCs differentiate into mature cells that initiate immune responses, while in the absence of infection, most of them remain in an immature form that induces tolerance to self Ags. Understanding what controls these opposing effects is an important goal for vaccine development and prevention of unwanted immune responses. A crucial question is what cytokine(s) regulates DC maturation in the absence of infection. In this study, we show that IL-6 plays a major role in maintaining immature DCs. IL-6 knockout (KO) mice had increased numbers of mature DCs, indicating that IL-6 blocks DC maturation in vivo. We examined this effect further in knockin mice expressing mutant versions of the IL-6 signal transducer gp130, with defective signaling through either Src homology region 2 domain-containing phosphatase 2/Gab/MAPK (gp130(F759/F759)) or STAT3 (gp130(FxxQ/FxxQ)), and combined gp130 and IL-6 defects (gp130(F759/F759)/IL-6 KO mice). Importantly, we found STAT3 activation by IL-6 was required for the suppression of LPS-induced DC maturation. In addition, STAT3 phosphorylation in DCs was regulated by IL-6 in vivo, and STAT3 was necessary for the IL-6 suppression of bone marrow-derived DC activation/maturation. DC-mediated T cell activation was enhanced in IL-6 KO mice and suppressed in gp130(F759/F759) mice. IL-6 is thus a potent regulator of DC differentiation in vivo, and IL-6-gp130-STAT3 signaling in DCs may represent a critical target for controlling T cell-mediated immune responses in vivo.     

脂肪细胞的内分泌和自分泌/旁分泌功能

脂肪细胞是高度特化的细胞,除了可通过内分泌、旁分泌和自分泌方式来调节脂肪组织本身的能量储存和释放外,还可以通过循环系统调控其他组织器官的代谢活动。     

p42/p44-MAPK and PI3K are sufficient for IL-6 family cytokines/gp130 to signal to hypertrophy and survival in cardiomyocytes in the absence of JAK/STAT activation

The effect of differential signalling by IL-6 and leukaemia inhibitory factor (LIF) which signal by gp130 homodimerisation or LIFRβ/gp130 heterodimerisation on survival and hypertrophy was studied in neonatal rat cardiomyocytes. Both LIF and IL-6 [in the absence of soluble IL-6 receptor (sIL-6Rα)] activated Erk1/2, JNK1/2, p38-MAPK and PI3K signalling peaking at 20 min and induced cytoprotection against simulated ischemia-reperfusion injury which was blocked by the MEK1/2 inhibitor PD98059 but not the p38-MAPK inhibitor SB203580. In the absence of sIL-6R, IL-6 did not induce STAT1/3 phosphorylation, whereas IL-6/sIL-6R and LIF induced STAT1 and STAT3 phosphorylation. Furthermore, IL-6/sIL-6R induced phosphorylation of STAT1 Tyr⁷⁰¹ and STAT3 Tyr⁷⁰⁵ were enhanced by SB203580. IL-6 and pheneylephrine (PE), but not LIF, induced cardiomyocyte iNOS expression and nitric oxide (NO) production. IL-6, LIF and PE induced cardiomyocyte hypertrophy, but with phenotypic differences in ANF and SERCA2 expression and myofilament organisation with IL-6 more resembling PE than LIF. Transfection of cardiomyocytes with full length or truncated chimaeric gp130 cytoplasmic domain/Erythropoietin receptor (EpoR) extracellular domain fusion constructs showed that the membrane proximal Box 1 and Box 2 containing region of gp130 was necessary and sufficient for MAPK and PI3K activation; hypertrophy; SERCA2 expression and iNOS/NO induction in the absence of JAK/STAT activation. In conclusion, IL-6 can signal in cardiomyocytes independent of sIL-6R and STAT1/3 and furthermore, that Erk1/2 and PI3K activation by IL-6 are both necessary and sufficient for induced cardioprotection. In addition, p38-MAPK may act as a negative feedback regulator of JAK/STAT activation in cardiomyocytes.     

IL-6/STAT3报告基因系统的构建及抑制IL-6/STAT3信号通路的中药单体的筛选及验证

寻找可抑制IL-6/STAT3信号通路的活化从而抑制肿瘤的生长和恶化的中药单体化合物具有重要意义及发展前景。文中通过基因重组技术构建出一种含有STAT3增强子序列和NanoLuc(NLuc)报告基因序列的新表达载体,并进一步建立受STAT3调控并稳定表达NLuc荧光素酶的细胞系,利用该细胞系定量检测多种中药单体化合物对IL-6/STAT3信号通路的调控作用,并对抑制IL-6/STAT3信号通路的中药单体的效果进行验证。酶切鉴定及测序结果表明报告基因表达载体pQCXIP-STAT3-NLuc构建成功。STAT3转录因子的刺激物白细胞介素-6(IL-6)作用于所构建的稳定表达NLuc的细胞系后出现特异性荧光素酶反应,且作用效果呈良好的剂量依赖性,表明受STAT3调控稳定表达NLuc荧光素酶的细胞系构建成功。Western blotting及Real-time PCR实验结果表明所筛选的中药单体化合物石斛碱及粉防己碱可抑制IL-6/STAT3信号通路并显著下调其下游基因Bcl-2及Bcl-x的表达,且作用呈剂量依赖性。综上所述,文中构建了可高效检测STAT3转录活性的报告基因系统,并利用该系统成功地筛选出可抑制IL-6/STAT3信号通路的中药单体化合物,具有一定的理论和应用价值。     

STAT3 mutations correlated with hyper-IgE syndrome lead to blockage of IL-6/STAT3 signalling pathway

Of all the causes identified for the disease hyper-immunoglobulinemia E syndrome (HIES), a homozygous mutation in tyrosine kinase2 (TYK2) and heterozygous mutations in STAT3 are implicated the defects in Jak/STAT signalling pathway in the pathogenesis of HIES. Mutations of STAT3 have been frequently clinically identified in autosomaldominant (AD) HIES patients' cells, and therefore, the genotype of STAT3 has been associated with the phenotype of HIES. Here, we conducted studies on the functional loss of the seven specific STAT3 mutations correlated with ADHIES. Using STAT3-null human colon carcinoma cell line A4 cells, we generated seven mutants of STAT3 bearing single mutations clinically identified in AD-HIES patients' cells and studied the functional loss of these mutants in IL- 6-Jak/STAT3 signalling pathway. Our results show that five STAT3 mutants bearing mutations in the DNA-binding domain maintain the phosphorylation of Tyr705 and the ability of dimerization while the other two with mutations in SH2 domain are devoid of the phosphorylation of Try705 and abrogate the dimerization in response to IL-6. The phosphorylation of Ser727 in these mutants shows diversity in response to IL-6. These mutations eventually converge on the abnormalities of the IL-6/Gp130/Jak2-mediated STAT3 transactivation on target genes, indicative of the dysregulation of JAK/STAT signalling present in HIES.     

IL-10 family of cytokines

In 2001, six immune mediators (IL-10, IL-19, IL-20, IL-22, IL-24, and IL-26) were grouped into the so-called IL-10 family of cytokines based on their similarities with respect to the structure and location of their encoding genes, their primary and secondary protein structures, and the receptor complexes used. Surprisingly, despite all these similarities, IL-10 family members possess different biological functions. The currently known facts regarding the biological effects of these six immune mediators give the impression that at least IL-10, IL-20, and IL-22 play an important role in the pathogenesis of some chronic inflammatory diseases. This review provides an overview of the most important and common aspects of the IL-10 family members.