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Retinoic acid receptor-related orphan receptor alpha (RORα), an essential tumor suppressor in a range of human malignancies, is classified as a member of the orphan nuclear receptor family. The most prevalent form of oral cancer, tongue squamous cell carcinoma (TSCC) is characterized by its severe malignancy and unfavorable prognosis. However, the extent to which its tumorigenesis mechanisms are associated with RORα expression levels is still not fully understood. The objective of this study was to examine the molecular mechanisms by which RORα is involved in TSCC. Through the use of immunohistochemistry (IHC), it was discovered that the expression level of RORα was significantly downregulated in TSCC tissues when compared to adjacent normal tissues in this study. To further investigate the role of RORα in TSCC, we activated the expression of RORα in human TSCC cell line (SCC9 cells) by transfecting RORα cDNA and using the selective RORα agonist SR1078. The results show that RORα can significantly inhibit the invasion, migration, proliferation, and adhesion of TSCC cells and induce cell apoptosis. In addition, xenograft models confirmed the conclusion that stable activation or treatment with SR1078 to increase RORα content significantly inhibited tumor growth and development. Taken together, this study provides solid evidence for the inhibitory role of RORα in the progression of TSCC. In addition, the preliminary application results of SR1078 in TSCC show that SR1078 is expected to be a potential therapeutic medication for TSCC. These findings provide innovative perspectives on the development of potential biomarkers and agents for TSCC therapy. The objective is to introduce novel strategy and alternatives for the prevention and treatment of TSCC. • RORα is an essential factor in the development and progression of TSCC. • RORα potentially serves a pivotal function in inhibiting the malignant attributes of TSCC. TSCC is inhibited by synthetic SR1078 via regulation of RORα expression. • RORα has the potential to serve as a valuable biomarker in the diagnosis and treatment of TSCC. Therapeutic potential was demonstrated for SR1078 as an anticancer medication. [ABSTRACT FROM AUTHOR]

Systemic inflammatory response syndrome (SIRS) is associated with excessive inflammatory response, however, the pathophysiology of inflammation is poorly understood. The retinoid-related orphan receptor α (RORα) is a key inflammatory regulator, but the mechanisms underlying its role remain unclear. The aim of this study was to investigate how RORα was involved in the regulation of inflammatory response. Here we put forward a hypothesis that RORα might negatively regulate inflammatory response by controlling silent information regulator Sirtuin 1 (SIRT1) expression. Stimulation of macrophages in vitro with LPS and LPS administration in vivo were used to explore the function of RORα and the relationship between RORα and SIRT1. We found that the level of RORα was suppressed in macrophages stimulated with LPS and overexpression or knockdown of RORα by transfection with lentivirus or siRNAs significantly decreased or increased, respectively, the pro-inflammatory cytokines IL-1β, TNF, IL-6 and MCP-1. Importantly, overexpression of RORα suppressed inflammation and alleviated LPS-induced organ injury in vivo. Further study showed that RORα could regulate SIRT1 expression and, consequently, affect deacetyation and nuclear translocation of nuclear factor-kappa B (NF-κB) subunit p65. Moreover, the activation of SIRT1 by its specific agonist, SR1720, could reduce the expression of proinflammatory cytokines in RORα knockdown macrophages stimulated with LPS. In conclusion, we demonstrated that RORα could alleviate LPS-induced inflammation and organ injury both in vivo and in vitro by blocking NF-κB p65 nuclear translocation and restricting acetylation of NF-κB p65 at lysine 310 via the regulation of SIRT1 expression. Targeting RORα might be a promising therapeutic strategy to regulate inflammatory disorders. [ABSTRACT FROM AUTHOR]

Hepatic gluconeogenic programs are regulated by a variety of signaling cascades. Glucagon – cAMP signaling is the main initiator of the gluconeogenic programs, including glucose-6-phosphatase catalytic subunit ( G6pc ) gene expression. Pterosin B, an ingredient in Pteridium aquilinum , inhibits salt-inducible kinase 3 signaling that represses cAMP-response element-binding protein regulated transcription coactivator 2, an inducer of gluconeogenic programs. As the results, pterosin B promotes G6pc expression even in the absence of cAMP. In this work, however, we noticed that once cAMP signaling was initiated, pterosin B became a strong repressor of G6pc expression. The search for associated transcription factors for pterosin B actions revealed that retinoic acid receptor-related orphan receptor alpha–steroid receptor coactivator 2 (RORα–SRC2) complex on the G6pc promoter was the target. Meanwhile, pterosin B impaired the oxidation–reduction cycle of coenzyme Q in mitochondrial oxidative phosphorylation (OXPHOS); and antimycin A, an inhibitor of coenzyme Q: cytochrome c–oxidoreductase (termed mitochondrial complex III), also mimicked pterosin B actions on RORα–SRC2 signaling. Although other respiratory toxins (rotenone and oligomycin) also suppressed G6pc expression accompanied by lowered ATP levels following the activation of AMP-activated kinase, minimal or no effect of these other toxins on RORα–SRC2 activity was observed. These results suggested that individual components in OXPHOS differentially linked to different transcriptional machineries for hepatic gluconeogenic programs, and the RORα–SRC2 complex acted as a sensor for oxidation–reduction cycle of coenzyme Q and regulated G6Pc expression. This was a site disrupted by pterosin B in gluconeogenic programs. [ABSTRACT FROM AUTHOR]

Maresin-1 is one of the representative specialized pro-resolving mediators that has shown beneficial effects in inflammatory disease models. Recently, two distinct types of receptor molecules were discovered as the targets of maresin-1, further revealing the pro-resolution mechanism of maresin-1. One is retinoic acid-related orphan receptor α (RORα) and the another one is leucine-rich repeat domain-containing G protein-coupled receptor 6 (LGR6). In this review, we summarized the detailed role of maresin-1 and its two different receptors in respiratory diseases. RORα and LGR6 are potential targets for the treatment of respiratory diseases. Future basic research and clinical trials on MaR1 and its receptors should provide useful information for the treatment of respiratory diseases. [ABSTRACT FROM AUTHOR]

Abstract: Purkinje cells (PCs) provide the primary output from the cerebellar cortex, which controls movement and posture, and loss of PCs causes severe cerebellar dysfunction. The mechanisms underlying cell fate determination and early differentiation of PC remain largely unknown. Here we show that the c-Ski family member and transcriptional regulator Corl2 is required for correct differentiation of PCs. In Corl2 knock-out embryos, initial PC specification appeared largely normal, but in a subset of presumptive PCs generated near the ventral border of the PC domain, cell fate choice was compromised and cells showed a mixed identity expressing the interneuron marker Pax2 as well. Additionally, selection and maintenance of the transmitter phenotype was compromised in most developing PCs in the mutants. During later differentiation steps, induction of PC marker genes was significantly suppressed, suggesting that maturation was delayed in the absence of Corl2. Consistently, defects in migration, cell polarization and dendrite formation were observed in mutant PCs, although their axonal trajectories appeared normal. These phenotypes closely resembled those of mutants for Rora, an essential regulator of PC differentiation. However, Rora expression was not significantly changed in the Corl2 mutants, indicating that Corl2 does not simply act upstream of Rora to promote PC differentiation. ChIP experiments revealed that Corl2 bound to the promoter regions of several PC-selective genes, which are also known to be direct downstream targets of RORα. Altogether, our results identified a novel regulatory program of PC differentiation involving Corl2, which might cooperate with the RORα pathway. [Copyright &y& Elsevier]

Abstract: Cholesterol sulfate is abundant in the human epidermis and is a putative natural ligand for retinoic acid receptor-related orphan receptor alpha (RORα). Although direct binding of cholesterol sulfate is expected to activate RORα, cholesterol sulfate can also induce RORα expression and increase RORα target gene expression. The purpose of this study was to determine whether cholesterol sulfate induces profilaggrin expression, a precursor of the barrier protein filaggrin in the epidermis, through activation of RORα by directly binding to RORα, or through increased RORα expression. Immunohistochemical and polymerase chain reaction (PCR) analyses showed that RORα was expressed in normal human epidermal keratinocytes (NHEKs) and that its expression increased during keratinocyte differentiation in parallel with that of profilaggrin and cholesterol sulfotransferase, which catalyzes the synthesis of cholesterol sulfate. Exogenous cholesterol sulfate significantly increased both RORα and profilaggrin expression in NHEKs, whereas no effect on profilaggrin expression was observed in cells in which RORα was knocked down with small interfering RNA (siRNA). Additionally, a luciferase reporter gene assay revealed that exogenous RORα dose-dependently increased the activity of the profilaggrin gene promoter even in the absence of cholesterol sulfate, and that this response involves activator protein-1. In conclusion, the results of this study indicate that cholesterol sulfate induces filaggrin expression through increased RORα expression. Further studies are required to fully elucidate the mechanisms involved. [Copyright &y& Elsevier]

Abstract: The retinoic acid related orphan receptor RORα positively regulates the transcription of genes important for cerebellar development, immune function, lipid metabolism, and circadian rhythm. In the present study, we identified protein kinase A (PKA) as RORα4 phosphorylating kinase in vitro. The primary sequence of RORα4 contains a PKA recognition motif (R-D-S99) within the c-terminal extension of the DNA-binding domain, and mutation of Ser-99 to Ala prevents RORα4 phosphorylation by PKA. Activation of PKA by dBcAMP results in a marked induction of RORα4 activity. Inhibition of PKA with the selective kinase inhibitor H89 inhibits dBcAMP mediated as well as CaMK-IV triggered increase in RORα4 transcriptional activity. The regulation of RORα activity by PKA as well as CaMK-IV provides a new link in the signalling network that regulates metabolic processes such as glycogen and lipid metabolism. [Copyright &y& Elsevier]

Abstract: SUMOylation regulates a variety of cellular processes, including control of transcriptional activities of nuclear receptors. Here, we present SUMOylation of orphan nuclear receptor, RORα by both SUMO-1 and SUMO-2. SUMOylation of RORα occurred on the 240th lysine residue at the hinge region of human protein. PIAS family members, PIASxα, PIAS3, and PIASy, increased SUMOylation of RORα, whereas SENP2 specifically removed SUMO from RORα. SUMOylation-defective mutant form of RORα exhibited decreased transcriptional activity on RORα-responsive promoters indicating that SUMOylation may positively regulate transcriptional function of RORα. [Copyright &y& Elsevier]

Abstract: The retinoic acid related orphan receptor RORα activates transcription of genes that play an important role in cerebellar development, the protection against age-related degenerative processes, the regulation of inflammatory responses, and is one of the pivotal participants that control the circadian rhythmicity in the core-clock of mammals. We identified the extracellular signal-regulated kinase 2 (ERK-2) as RORα4 phosphorylating kinase in vitro. The primary sequence of RORα4 contains an ERK-2 recognition motif (P-L-T128-P) within the hinge domain, and mutation of Thr-128 to Ala prevents RORα4 phosphorylation by ERK. The RORα4-T128A mutant exhibits an increased DNA-binding affinity, an increased transcriptional activity and, in the interplay with the opponent RevErbα, acts as a stronger competitor at ROR response elements than RORα4-WT. [Copyright &y& Elsevier]

Abstract: RORα4 is a nuclear receptor activating the transcription of genes that are important for a variety of physiological processes like muscle differentiation, lipid and bone metabolism, cerebellar development, and inflammation. Furthermore, it plays an essential role in maintaining circadian rhythmicity of the core clock in the suprachiasmatic nuclei (SCN). Here, we describe the successful overexpression and purification of human full-length RORα4 in Escherichia coli using a T7 expression system. The expressed protein formed inclusion bodies which were solubilized in the presence of 6M guanidinium–HCl and renatured by gradual removal of guanidinium–HCl and addition of l-arginine. The refolded protein was purified by nickel affinity chromatography due to an N-terminal polyhistidine tag which can be cleaved with thrombin subsequently. This method permitted us to obtain up to 20mg of pure and native RORα4 protein per liter of E. coli culture. The DNA binding activity of the refolded protein was demonstrated by electrophoretic mobility shift assay (EMSA) using an oligonucleotide comprising the ROR-response element (RORE) motif (A/G)GGTCA. In addition, we developed a new monoclonal antibody to human RORα in mice with high sensitivity and specificity. [Copyright &y& Elsevier]

Abstract: Retinoic acid receptor-related orphan receptor-α (RORα) (NR1F1) is an orphan nuclear receptor with a potential role in metabolism. Previous studies have shown that RORα regulates transcription of the murine Apolipoprotein AI gene and human Apolipoprotein CIII genes. In the present study, we present evidence that RORα also induces transcription of the human Apolipoprotein AV gene, a recently identified apolipoprotein associated with triglyceride levels. Adenovirus-mediated overexpression of RORα increased the endogenous expression of ApoAV in HepG2 cells and RORα also enhanced the activity of an ApoAV promoter construct in transiently transfected HepG2 cells. Deletion and mutation studies identified three AGGTCA motifs in the ApoAV promoter that mediate RORα transactivation, one of which overlaps with a previously identified binding site for PPARα. Together, these results suggest a novel mechanism whereby RORα modulates lipid metabolism and implies RORα as a potential target for the treatment of dyslipidemia and atherosclerosis. [Copyright &y& Elsevier]

The ligand-binding domain (LBD) of the human retinoic acid receptor-related orphan receptor (RORα-LBD), expressed in Sf9 cells, was purified and analyzed by electrospray ionization–mass spectrometry (ESI-MS). ESI-MS operated under native conditions showed the presence of a fortuitous ligand with molecular weight 386. Further analysis by gas chromatography–mass spectrometry (GC-MS) allowed the identification of the ligands bound to the LBD. Cholesterol (77%) and 7-dehydrocholesterol (provitamin D3; 18%) were shown to be the major ligands. A monohydroxylated cholesterol derivative was identified as a minor ligand. In addition, ligand exchange experiments monitored by ESI-MS showed that cholesterol sulfate has a higher affinity for RORα-LBD than cholesterol and 25-hydroxycholesterol. Binding of coactivator (CoA) peptide GRIP1P was shown to occur in a stoichiometric manner. Therefore, monitoring of binding of CoAs by mass spectrometry could be used for classification of the ligands as agonist or antagonist molecules. [Copyright &y& Elsevier]