Cloning and characterization of the human beta2-glycoprotein I (beta2-GPI) gene promoter: roles of the atypical TATA box and hepatic nuclear factor-1alpha in regulating beta2-GPI promoter activity.
Beta2-glycoprotein I (beta2-GPI) is a plasma glycoprotein primarily synthesized in the liver. The interindividual variability of beta2-GPI expression in subjects with various metabolic syndromes and disease states suggests that it may have clinical importance. However, the regulation of beta2-GPI gene expression has not been clarified. To gain more insight into the control of beta2-GPI gene expression, we cloned the 4.1-kb 5'-flanking region and characterized the proximal promoter of the beta2- GPI gene in this study. Cis -acting elements required for beta2-GPI promoter activity were identified with transient transfection assays in the hepatoma cell lines HepG2 and Huh7 and in non-hepatic HeLa cells. Serial deletion analyses of the beta2-GPI 5'-flanking sequence revealed that the region from -197 to +7 had strong promoter activity in hepatoma cells but not in HeLa cells. Truncation and site-directed mutagenesis of putative cis -elements within this region showing an atypical TATA box and a HNF-1 (hepatic nuclear factor-1) element were both essential for the beta2-GPI promoter activity. Subsequent gel mobility shift assays confirmed the interaction of HNF-1alpha with the HNF-1 site residing downstream of the TATA box. Co-transfection of beta2-GPI promoter-luciferase vector with HNF-1alpha expression vector in Huh7 and HNF-1-deficient HeLa cells demonstrated the transactivation effect of HNF-1alpha on beta2-GPI promoter activity. In addition, overexpression of HNF-1alpha enhanced the endogenous beta2-GPI expression. These results suggest that the atypical TATA box and HNF-1 cis-element are critical for beta2-GPI transcription and HNF-1alpha may play an important role in cell-specific regulation of beta2-GPI gene expression.
Institute of Biochemistry, School of Life Science, National Yang-Ming University, Taipei 112, Taiwan, ROC.
Hsueh-Hsiao
HH
Wang
An-Na
AN
Chiang
D024506:5' Flanking Region / Q000235:genetics; D001483:Base Sequence; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000473:pathology; D045744:Cell Line, Tumor; D003001:Cloning, Molecular / Q000379:methods*; D004273:DNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D004742:Enhancer Elements, Genetic / Q000235:genetics; D006023:Glycoproteins / Q000235:genetics*; D006367:HeLa Cells / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000473:pathology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D016385:TATA Box / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism; D024363:Transcription Initiation Site; D015533:Transcriptional Activation / Q000235:genetics; D014162:Transfection / Q000379:methods; D053482:beta 2-Glycoprotein I
D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D006023:Glycoproteins; C496487:HNF1A protein, human; D051538:Hepatocyte Nuclear Factor 1-alpha; D009687:Nuclear Proteins; D014157:Transcription Factors; D053482:beta 2-Glycoprotein I; D051537:Hepatocyte Nuclear Factor 1