title: Promoter characterization and role of cAMP/PKA/CREB in the basal transcription of the mouse ORMDL3 gene.
abstract: Orosomucoid 1-like 3 (ORMDL3) gene was strongly linked with the development of asthma in genetic association studies, and its expression could be significantly induced by allergen in airway epithelial cells of mice. However, the expression mechanism of ORMDL3 was still unclear. Here we have identified and characterized the mouse ORMDL3 gene promoter. Deletion constructs of the 5' ﬂanking region were fused to a luciferase reporter gene. After transient transfection in mouse fibroblast cell line NIH3T3, a CRE (-27/-20) binding CREB was identified in the core promoter region. Deletion or mutation of the CRE consensus sequence resulted in a significant loss of the promoter activity. EMSA and ChIP assays demonstrated the binding of CREB to the core promoter. Knocking down endogenous CREB led to a reduction in ORMDL3 expression. Conversely, overexpression of CREB up-regulated ORMDL3 expression. Moreover, forskolin, a PKA activator, could facilitate the phosphorylation of CREB, which in turn heightens ORMDL3 expression. H-89, a PKA-specific inhibitor, could significantly inhibit ORMDL3 expression. This study delineates the characterization of mouse ORMDL3 gene promoter and shows signaling pathway cAMP/PKA/CREB plays an important role in regulating ORMDL3 expression, which will be helpful for future animal model studies regarding the regulation or function of ORMDL3 gene.
authors.affiliation: Department of Pediatrics, The First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province, China.
authors.forename: Li-Li
authors.initials: LL
authors.lastname: Zhuang
authors.forename: Rui
authors.initials: R
authors.lastname: Jin
authors.forename: Liang-Hua
authors.initials: LH
authors.lastname: Zhu
authors.forename: Hua-Guo
authors.initials: HG
authors.lastname: Xu
authors.forename: Yue
authors.initials: Y
authors.lastname: Li
authors.forename: Shan
authors.initials: S
authors.lastname: Gao
authors.forename: Jia-Yin
authors.initials: JY
authors.lastname: Liu
authors.forename: Guo-Ping
authors.initials: GP
authors.lastname: Zhou
mesh_terms: D000818:Animals; D001481:Basal Metabolism; D001483:Base Sequence; D000242:Cyclic AMP / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000235:genetics; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D020218:Response Elements / Q000235:genetics; D015398:Signal Transduction / Q000235:genetics; D014158:Transcription, Genetic
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D008565:Membrane Proteins; C583840:ORMDL3 protein, mouse; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases