Signal transducer and activator of transcription 6 directly regulates human ORMDL3 expression

• Published in: The FEBS journal 2013-05, Vol.280 (9), p.2014-2026
• Main Authors: Qiu, Rongfang, Yang, Yang, Zhao, Hailing, Li, Jiangxia, Xin, Qian, Shan, Shan, Liu, Yongchao, Dang, Jie, Yu, Xiao, Gong, Yaoqin, Liu, Qiji
• Format: Article
• Language: English
• Subjects: Base Sequence; Binding Sites; Cell Line, Tumor; Consensus Sequence; Gene expression; Gene Expression Regulation; Gene Knockdown Techniques; Genes, Reporter; Genomics; HEK293 Cells; Humans; Inflammatory diseases; Interleukin-13 - physiology; Interleukin-4 - physiology; Leukocytes, Mononuclear - metabolism; Membrane Proteins - genetics; Membrane Proteins - metabolism; ORMDL3; p300; Promoter Regions, Genetic; Protein Binding; Signal transduction; STAT6; STAT6 Transcription Factor - genetics; STAT6 Transcription Factor - metabolism; STAT6 Transcription Factor - physiology; Th2 cytokines; transcription regulation; Transcriptional Activation
• ISSN: 1742-464X; 1742-4658
• DOI: 10.1111/febs.12225

Orosomucoid‐like 3 (ORMDL3) has been associated with asthma and a series of autoimmune disorders, and is involved in endoplasmic reticulum‐mediated inflammatory responses. However, its clinical significance and the molecular mechanism underlying its expression are still largely unclear. To elucidate the mechanisms of human ORMDL3 transcriptional regulation, we cloned a 1.5 kb genomic DNA fragment containing the putative promoter region and evaluated its transcriptional activity in a luciferase reporter system by deletion analysis. We identified a 68 bp region that functions as a minimal promoter. Bioinformatics analysis predicted that the −64 to −56 bp region contained a signal transducer and activator of transcription 6 (STAT6) binding site. Electrophoretic mobility shift assay and chromatin immunoprecipitation demonstrated that STAT6 bound to its binding site within the ORMDL3 promoter. STAT6 over‐expression or knockdown trans‐activated or trans‐inhibited, respectively, the ORMDL3 promoter containing the STAT6‐binding motif. Treatment with interleukins 4 or 13 increased ORMDL3 promoter activity as well as endogenous ORMDL3 expression. Immunoprecipitation and ChIP/Re‐ChIP assays revealed that STAT6 and p300 exist in the same protein complex that binds to the ORMDL3 promoter. Our study confirmed that STAT6 plays important roles in regulating the expression of human ORMDL3 by directly binding to the promoter region, which may shed light on a possible role in various human diseases. Structured digital p300 physically interacts with STAT6 by anti bait coimmunoprecipitation (View Interaction: 1, 2) To elucidate the mechanisms of ORMDL3 transcriptional regulation, we cloned the 1.5‐kb genomic DNA fragment and identified a 68 bp region that functions as a minimal promoter. EMSA, ChIP and ChIP/Re‐ChIP assays revealed that STAT6 and p300 exist in the same protein complex binding to the ORMDL3 promoter. Our study confirmed that STAT6 plays a role in regulating human ORMDL3.