Identification of the promoter region required for human adiponectin gene transcription: Association with CCAAT/enhancer binding protein-b and tumor necrosis factor-a

Adiponectin. an adipose tissuee-specific plasma protein. is involved in insulin Sensitizing and has anti-atherosclerotic properties. Plasma levels of adiponectin are decreased in obese individuals and patients with type 2 diabetes with insulin resistance, Tumor necrosis factor-a (TNF-a) decreases the expression of adiponectin in adipocytes, The aims of the present study were: (1) to identify the promoter region responsible for basal transcription of the human adiponectin gene. and (2) to investigate the mechanism by which adiponectin was regulated by TNF-a. The human adiponectin pronioter (2.1 kb) as isolated and used for luciferase reporter analysis by transient transfection into 3T3-LI adipocytes. Deletion analysis demonstrated that the promoter region from -676 to +41 was sufficient for basal transcriptional activity. Mutation analysis of putative response elements for sterol regulatory element binding protein (SREBP) (-431 to -423) and CCAAT/enhancer binding protein (c/EBP) (- 230 to 224) showed that both elements were required for basal promoter activity. Adiponectin transcription was increased 3-fold in cells that over-expressed constitutively active C/EBP-b. Electrophoretic mobility shift assay, using nuclear extract front 3T3-L1 cells and the 258 to - 199 region as a probe, demonstrated specific DNA-protein binding, which was abolished by TNF-a treatment. The present data indicate that the putative response elements for SREBP and C/EBP are required Cor human adiponectin promoter activity, and that suppression by TNF-a may, at least in part, be associated with inactivation of C/EBP-b. 2005 Elsevier Inc. All rights reserved.