AT-rich Interactive Domain-containing Protein 5a Research Articles

Central Roles of STAT3-Mediated Signals in Onset and Development of Cancers: Tumorigenesis and Immunosurveillance.

Since the time of Rudolf Virchow in the 19th century, it has been well-known that cancer-associated inflammation contributes to tumor initiation and progression. However, it remains unclear whether a collapse of the balance between the antitumor immune response via the immunological surveillance system and protumor immunity due to cancer-related inflammation is responsible for cancer malignancy. The majority of inflammatory signals affect tumorigenesis by activating signal transducer and activation of transcription 3 (STAT3) and nuclear factor-κB. Persistent STAT3 activation in malignant cancer cells mediates extremely widespread functions, including cell growth, survival, angiogenesis, and invasion and contributes to an increase in inflammation-associated tumorigenesis. In addition, intracellular STAT3 activation in immune cells causes suppressive effects on antitumor immunity and leads to the differentiation and mobilization of immature myeloid-derived cells and tumor-associated macrophages. In many cancer types, STAT3 does not directly rely on its activation by oncogenic mutations but has important oncogenic and malignant transformation-associated functions in both cancer and stromal cells in the tumor microenvironment (TME). We have reported a series of studies aiming towards understanding the molecular mechanisms underlying the proliferation of various types of tumors involving signal-transducing adaptor protein-2 as an adaptor molecule that modulates STAT3 activity, and we recently found that AT-rich interactive domain-containing protein 5a functions as an mRNA stabilizer that orchestrates an immunosuppressive TME in malignant mesenchymal tumors. In this review, we summarize recent advances in our understanding of the functional role of STAT3 in tumor progression and introduce novel molecular mechanisms of cancer development and malignant transformation involving STAT3 activation that we have identified to date. Finally, we discuss potential therapeutic strategies for cancer that target the signaling pathway to augment STAT3 activity.

Differential modulation of Ahr and Arid5a: A promising therapeutic strategy for autoimmune encephalomyelitis

Multiple sclerosis (MS) is an autoimmune disease that involves demyelination of axons in the central nervous system (CNS) and affects patients worldwide. It has been demonstrated that ligand-activated aryl hydrocarbon receptor (Ahr) ameliorates experimental autoimmune encephalomyelitis (EAE), a murine model of MS, by increasing CD4+FoxP3+ T cells. Recent evidence indicates that AT-rich interactive domain-containing protein 5a (Arid5a) is required for EAE pathogenesis by stabilizing Il6 and OX40 mRNAs. However, the differential modulation of Ahr and Arid5a in autoimmunity as a therapeutic strategy is unexplored. Herein, an in silico, in vitro and in vivo approach identified Flavipin (3,4,5-trihydroxy-6-methylphthalaldehyde) as an Ahr agonist that induces the expression of Ahr downstream genes in mouse CD4+ T cells and CD11b+ macrophages. Interestingly, Flavipin inhibited the stabilizing function of Arid5a and its counteracting effects on Regnase-1 on the 3′ untranslated region (3′UTR) of target mRNAs. Furthermore, it inhibited the stabilizing function of Arid5a on Il23a 3′UTR, a newly identified target mRNA. In EAE, Flavipin ameliorated disease severity, with reduced CD4+IL-17+ T cells, IL-6 and TNF-α and increased CD4+FoxP3+ T cells. Moreover, EAE amelioration was concomitant with reduced CD4+OX40+ and CD4+CD45+ T cells in the CNS. RNA interference showed that the modulatory effects of Flavipin on pro- and anti-inflammatory mediators in CD4+ T cells and macrophages were Ahr- and/or Arid5a-dependent. In conclusion, our findings reveal differential modulation of Ahr and Arid5a as a new therapeutic strategy for MS.

Differential effects of Flavipin on Ahr and Arid5a: A new therapeutic strategy for autoimmune encephalomyelitis

Abstract Multiple sclerosis (MS) is a World-wide spread autoimmune disease that includes episode of demyelination in the central nervous system (CNS). Accumulating evidence has demonstrated that ligand-activated aryl hydrocarbon receptor (Ahr) improves pathological outcomes in experimental autoimmune encephalomyelitis (EAE), an experimental model of MS. Recent evidences have shown that AT-rich interactive domain-containing protein 5a (Arid5a), an RNA-binding protein, is required for development of EAE via stabilizing the mRNAs of interleukin-6 (Il6) and Ox40. To date, there are no reports studying both Ahr and Arid5a as a potential therapeutic strategy for MS. Thus, current work applied in silico, in vitro and in vivo approaches to identify a molecule that interacts with Ahr and Arid5a. The results identified Flavipin (3,4,5-trihydroxy-6-methylphthalaldehyde) as a new Ahr agonist that induces the expression of downstream genes in mice CD4+ T cells and CD11b+ macrophages. Interestingly, Flavipin inhibited the stabilizing function of Arid5a, and impaired its counteracting effects on RNase function of Regnase-1 on 3′ untranslated region (3′UTR) of Il6, Ox40 and signal transducer and activator of transcription 3 (stat3). In EAE, Flavipin ameliorated disease symptoms concomitant with inducing acetylcholinesterase (AChE)-targeting miR-132. It reduced the expression of Ox40 in CD4+ T cells and the number infiltrated CD4+CD45+ T cells in the CNS. Further analysis identified Il23a mRNA as a new target for Arid5a. Taken together, our findings provide first evidences on differential effects of Flavipin on Ahr and Arid5a and pave an avenue for a new therapeutic strategy for autoimmune diseases such as MS.

β2-adrenergic stimulation induces interleukin-6 by increasing Arid5a, a stabilizer of mRNA, through cAMP/PKA/CREB pathway in cardiac fibroblasts.

Background and PurposeIn cardiovascular diseases, cardiac fibroblasts (CFs) participate in the myocardial inflammation by producing pro‐inflammatory cytokines, worsening the prognosis. β2‐adrenergic receptor (AR) and β3AR are expressed in CFs, and β‐adrenergic stimulation promotes CFs to produce pro‐inflammatory cytokines. However, the mechanism of the expression of pro‐inflammatory cytokines in response to β‐adrenergic stimulation remains to be fully elucidated.Experimental ApproachCFs were isolated from adult wild‐type or AT‐rich interactive domain‐containing protein 5A (Arid5a) knockout mice. The expression of mRNA was measured by real‐time RT‐PCR. Interleukin (IL)‐6 protein was measured by ELISA. The activity of nuclear factor‐κB (NF‐κB) and cyclic AMP (cAMP) response element binding protein (CREB) was assessed by ELISA‐like assay or Western blotting.Key ResultsThe β‐adrenergic stimulation remarkably induced IL‐6 mRNA and protein through β2AR in CFs. The activation of adenylate cyclase and the enhancement of intracellular cAMP resulted in the upregulation of IL‐6 mRNA expression. The induction of IL‐6 transcript by β2AR signaling was independent of NF‐κB. Concomitant with IL‐6, the expression of Arid5a, an IL‐6 mRNA stabilizing factor, was enhanced by β2‐adrenergic stimulation and by cAMP increase. Importantly, β2AR signaling‐mediated IL‐6 induction was suppressed in Arid5a knockout CFs. Finally, β2AR stimulation phosphorylated CREB via PKA pathway, and the activation of CREB was essential for the induction of Arid5a and IL‐6 mRNA.Conclusion and Implicationsβ2‐adrenergic stimulation post‐transcriptionally upregulates the expression of IL‐6 by the induction of Arid5a through cAMP/PKA/CREB pathway in adult CFs. β2AR/Arid5a/IL‐6 axis could be a therapeutic target against cardiac inflammation.

Arid5a Regulation and the Roles of Arid5a in the Inflammatory Response and Disease.

Abnormal gene expression patterns underlie many diseases that represent major public health concerns and robust therapeutic challenges. Posttranscriptional gene regulation by RNA-binding proteins (RBPs) is well-recognized, and the biological functions of RBPs have been implicated in many diseases, such as autoimmune diseases, inflammatory diseases, and cancer. However, a complete understanding of the regulation mediated by several RBPs is lacking. During the past few years, a novel role of AT-rich interactive domain-containing protein 5a (Arid5a) as an RBP is being investigated in the field of immunology owing to binding of Arid5a protein to the 3′ untranslated region (UTR) of Il-6 mRNA. Indeed, Arid5a is a dynamic molecule because upon inflammation, it translocates to the cytoplasm and stabilizes a variety of inflammatory mRNA transcripts, including Il-6, Stat3, Ox40, T-bet, and IL-17-induced targets, and contributes to the inflammatory response and a variety of diseases. TLR4-activated NF-κB and MAPK pathways are involved in regulating Arid5a expression from synthesis to degradation, and even a slight alteration in these pathways can lead to intense production of inflammatory molecules, such as IL-6, which may further contribute to the development of inflammatory diseases such as sepsis and experimental autoimmune encephalomyelitis. This review highlights the regulation of the Arid5a expression and function. Additionally, recent findings on Arid5a are discussed to further our understanding of this molecule, which may be a promising therapeutic target for inflammatory diseases.

Arid5a orchestrates IL-17-mediated inflammation through post-transcriptional control of mRNA

Abstract Interleukin-17A (IL-17) is a proinflammatory cytokine essential for clearance of microbial pathogens, but conversely drives immunopathology in autoimmune settings. Surprisingly, very little is known about IL-17 receptor-mediated signaling pathway. Although IL-17 is consistently found to be a modest activator of gene transcription in experimental settings, its biological impact in vivo is profound. In addition to de novo transcription, IL-17 also upregulates inflammatory gene expression by stabilizing their transcripts. mRNA stabilization is a central, but still poorly understood facet of IL-17 signaling. Here, we report an RNA binding protein Arid5a (AT-rich interactive domain-containing protein 5A) as a novel mediator of IL-17-induced stabilization of inflammatory mRNAs. IL-17 stimulation upregulated Arid5a expression, whereupon Arid5a was rapidly recruited to TRAF2. Arid5a stabilized multiple IL-17-induced cytokine mRNA transcripts, including Il6, Cxcl1 and Cxcl5, by binding to their 3′ untranslated regions (UTR). In some cases, Arid5a counteracted mRNA degradation mediated by the endoribonuclease MCPIP1 (Regnase-1). Additionally, Arid5a enhanced IL-17-induced expression of the transcription factors IκBz (Nfkbiz) and C/EBPβ (Cebpb), which could then transactivate IL-17-dependent promoters. Arid5a bound to Nfkbiz or Cebpb transcripts, but surprisingly did not detectably affect total Nfkbiz or Cebpb mRNA levels. Rather, Arid5a facilitated translation of IkBz and C/EBPb. Thus, Arid5a orchestrates a feed-forward amplification loop within the IL-17 pathway, revealing a new function for Arid5a in controlling translation and demonstrating a role in controlling downstream IL-17 signaling.

Arid5a stabilizes OX40 mRNA in murine CD4+ T cells by recognizing a stem‐loop structure in its 3′UTR

AT-rich interactive domain-containing protein 5a (Arid5a) is an RNA-binding protein (RBP) required for autoimmunity via stabilization of interleukin-6 (Il6) and signal transducer and activator of transcription 3 (STAT3) mRNAs. However, the roles of Arid5a in Th17 cells and its association with autoimmunity remain unknown. Here, we show that the levels of Arid5a and OX40 are correlated in CD4+ Tcells under Th17 conditions in an IL-6-dependent manner. Lack of Arid5a in Tcells reduced OX40 expression levels and repressed IL-17 production in response to OX40 ligation. Arid5a stabilized OX40 mRNA by recognizing the alternative decay element (ADE)-like stem-loop (SL) in the 3' untranslated region (3'UTR). Interestingly, Arid5a impaired the RNA-destabilizing functions of Regnase-1 and Roquin-1 on OX40 ADE-like SL. In EAE, Arid5a-deficient mice exhibited resistance to EAE, with reduced OX40 expression in CD4+ Tcells, and the number of CD4+ CD45+ Tcells was decreased in CNS. Furthermore, ameliorated EAE was induced by adoptive transfer of Arid5a-/- encephalitogenic CD4+ Tcells expressing less OX40 mRNA and producing less IL-17. In conclusion, our findings indicate that the Arid5a/OX40 axis in CD4+ Tcells may have important implications in pathogenesis of autoimmune diseases such as EAE.

TLR4-induced NF-κB and MAPK signaling regulate the IL-6 mRNA stabilizing protein Arid5a.

The AT-rich interactive domain-containing protein 5a (Arid5a) plays a critical role in autoimmunity by regulating the half-life of Interleukin-6 (IL-6) mRNA. However, the signaling pathways underlying Arid5a-mediated regulation of IL-6 mRNA stability are largely uncharacterized. Here, we found that during the early phase of lipopolysaccharide (LPS) stimulation, NF-κB and an NF-κB-triggered IL-6-positive feedback loop activate Arid5a gene expression, increasing IL-6 expression via stabilization of the IL-6 mRNA. Subsequently, mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1) promotes translocation of AU-rich element RNA-binding protein 1 (AUF-1) from the nucleus to the cytoplasm, where it destabilizes Arid5a mRNA by binding to AU-rich elements in the 3΄ UTR. This results in downregulation of IL-6 mRNA expression. During the late phase of LPS stimulation, p38 MAPK phosphorylates Arid5a and recruits the WW domain containing E3 ubiquitin protein ligase 1 (WWP1) to its complex, which in turn ubiquitinates Arid5a in a K48-linked manner, leading to its degradation. Inhibition of Arid5a phosphorylation and degradation increases production of IL-6 mRNA. Thus, our data demonstrate that LPS-induced NF-κB and MAPK signaling are required to control the regulation of the IL-6 mRNA stabilizing molecule Arid5a. This study therefore substantially increases our understanding of the mechanisms by which IL-6 is regulated.

Arid5a regulates naive CD4+ T cell fate through selective stabilization of Stat3 mRNA.

Balance in signal transducer and activator of transcription (STAT) activation is a key factor in regulating the fate of naive CD4(+)T cells. Here, we demonstrate that AT-rich interactive domain-containing protein 5a (Arid5a) in T cells directs naive CD4(+)T cells to differentiate into inflammatory CD4(+)T cells, especially Th17 cells, through selective stabilization of Stat3(but not Stat1 and Stat5) mRNA in an IL-6-dependent manner. Loss of Arid5a in T cells led to reduction of STAT3 level under Th17-polarizing conditions, whereas STAT1 and STAT5 in Arid5a-deficient T cells were highly activated compared with those of WT T cells under the same conditions. These cells displayed the feature of antiinflammatory (Il10-expressing) CD4(+)T cells. Thus, we show a T cell-intrinsic role of Arid5a on fate decisions of naive CD4(+)T cells through selective stabilization of Stat3 mRNA.

AT-rich-interactive domain-containing protein 5A functions as a negative regulator of retinoic acid receptor-related orphan nuclear receptor γt-induced Th17 cell differentiation.

The proinflammatory cytokines tumor necrosis factor α and interleukin-6 (IL-6) and the Th17 cell cytokine IL-17A are implicated in the pathogenesis of rheumatoid arthritis (RA), and the blockade of these cytokines by biologic agents provides clinical benefits for RA patients. We undertook this study to clarify the mechanisms underlying the efficacy of IL-6 blockade in RA and to find a novel target for treatment of RA. We examined gene expression profiles of CD4+ T cells by DNA microarray analysis before and after treatment with an anti-IL-6 receptor antibody, tocilizumab (TCZ), in RA patients who exhibited good clinical responses to the treatment. Using murine CD4+ T cells, we then examined the roles of a newly identified molecule whose expression was significantly reduced in CD4+ T cells by TCZ therapy. We also examined the effect of the forced expression of the molecule on retinoic acid receptor-related orphan nuclear receptor γt (RORγt)-induced IL-17A production in CD4+ T cells and on RORγt-induced IL-17A promoter activation. We identified AT-rich-interactive domain- containing protein 5A (ARID-5A) as a new molecule down-regulated by IL-6 blockade in the form of TCZ therapy. IL-6 induced the expression of ARID-5A in CD4+ T cells during Th17 cell differentiation by a STAT-3-dependent mechanism, whereas IL-6-induced ARID-5A expression was not affected by the absence of RORγt, a lineage-specifying transcription factor of Th17 cells. Furthermore, ARID-5A physically associated with RORγt through its N-terminal region and inhibited RORγt-induced Th17 cell differentiation. ARID-5A is a lineage-specific attenuator of Th17 cell differentiation and may be involved in the pathogenesis of RA.

S-5: A novel IL-6 mRNA stability protein, Arid5A which is involved in IL-6 dependent autoimmunity

Deregulated expression of IL-6 is responsible for several chronic inflammations and hemopoietic malignancies. Humanized antibody against 80kd IL-6R (Tocilizumab) has shown significant therapeutic effect in RA, JIA, Castleman’s diseases, several other autoimmune inflammatory diseases, such as progressive sclerosis, reactive arthritis, polymyalgia rheumatica, adult still’s desease and uveoretinitis. Cytokine storm induced by hyperactivation of T cells has been shown to be controlled by Tocilizumab. An important question is how IL-6 production is regulated. Posttranscriptional regulation of IL-6 has been largely uncharacterized, with the exception of the ribonuclease Regnase-1, which prevents autoimmunity by destabilizing IL-6 mRNA. Here, we identified AT-rich interactive domain-containing protein 5A (Arid5a) as a unique RNA binding protein, which stabilizes IL-6 but not TNF- α mRNA through binding to the 3′ untranslated region of IL-6 mRNA. Arid5a was enhanced in macrophages in response to LPS, IL-1 β , and IL-6. Arid5a deficiency inhibited elevation of IL-6 serum level in LPS-treated mice and suppressed IL-6 levels and the development of T H 17 cells in experimental autoimmune encephalomyelitis. Importantly, Arid5a inhibited the destabilizing effect of Regnase-1 on IL-6 mRNA. These results indicate that Arid5a plays an important role in promotion of inflammatory processes and autoimmune diseases.

Arid5a controls IL-6 mRNA stability, which contributes to elevation of IL-6 level in vivo

Posttranscriptional regulation of IL-6 has been largely uncharacterized, with the exception of the ribonuclease Regnase-1, which prevents autoimmunity by destabilizing IL-6 mRNA. Here, we identified AT-rich interactive domain-containing protein 5A (Arid5a) as a unique RNA binding protein, which stabilizes IL-6 but not TNF-α mRNA through binding to the 3' untranslated region of IL-6 mRNA. Arid5a was enhanced in macrophages in response to LPS, IL-1β, and IL-6. Arid5a deficiency inhibited elevation of IL-6 serum level in LPS-treated mice and suppressed IL-6 levels and the development of T(H)17 cells in experimental autoimmune encephalomyelitis. Importantly, Arid5a inhibited the destabilizing effect of Regnase-1 on IL-6 mRNA. These results indicate that Arid5a plays an important role in promotion of inflammatory processes and autoimmune diseases.