Identification and characterization of a glucose-responsiveness region upstream of human insulin gene in transfected HIT-T 15 cells.
To determine possible regulation of full-length human insulin gene promoter activity by glucose, we examined a 2-kilobase pair (kbp) 5'-flanking region of the human insulin gene and characterized the DNA elements in transfected HIT-T 15 cells. The expression of the 2-kilobase pair 5'-flanking region human insulin gene fused to the luciferase reporter gene occurred by transfection. In 0.8 mM glucose of the F-12 K medium, the element mediating the negative regulatory region was localized from -1782 to -1295 base pairs (bp) and stimulatory element from -1295 to -1138 bp. The elements from -1138 to -880 bp and from -356 to +252 bp possessed the elements dose-dependently responsive to 0.8 mM, 7.0 and 22.2 mM glucose. In fragment D, cotransfection of oligonucleotide that confers RIPE3b1 activator decreased the glucose-stimulated promoter activity, but the other oligonucleotide that confers STF-1 did not. The present data indicated that 2 kbp possesses glucose-responsive region in the element from -1138 to -880 bp, in addition to the previously reported element from -356 to initiation site. There may exist a RIPE3b1 activator binding site in the glucose-responsive element from -1138 to -880 bp. In addition, negatively regulatory region may exist from -1782 to -1295 bp.
Ohtani
K
K
First Department of Internal Medicine, Gunma University School of Medicine, Japan.
Shimizu
H
H
Kato
Y
Y
Mori
M
M
D001665:Binding Sites; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D005947:Glucose / Q000494:pharmacology*; D018398:Homeodomain Proteins; D006801:Humans; D007328:Insulin / Q000235:genetics*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014162:Transfection / Q000235:genetics
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D007328:Insulin; D009838:Oligodeoxyribonucleotides; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein; D008156:Luciferases; D005947:Glucose