Characterization of a novel protein kinase C response element in the glucagon gene.
To maintain glucose levels in blood within narrow limits, the synthesis and secretion of pancreatic islet hormones are controlled by a variety of neural, hormonal, and metabolic messengers that act through multiple signal transduction pathways. Glucagon gene transcription is stimulated by cyclic AMP and depolarization-induced calcium influx. In this study, the effect of protein kinase C on glucagon gene transcription was investigated. After transient transfection of a glucagon-reporter fusion gene into the glucagon-producing islet cell line alphaTC2, activation of protein kinase C by 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulated glucagon gene transcription. By 5' deletions, 3' deletions, internal deletion, and oligonucleotide cassette insertion, the TPA-responsive element was mapped to the G2 element (from -165 to -200). Like TPA, overexpression of oncogenic Ras (V-12 Ras) stimulated G2-mediated transcription whereas overexpression of a dominant negative Ras mutant (N-17 Ras) blocked the effect of TPA. A mutational analysis of G2 function and nuclear protein binding indicated that protein kinase C and Ras responsiveness is conferred to the glucagon gene by HNF-3beta functionally interacting with a protein that binds to a closely associated site with sequence similarity to binding sites of Ets family proteins. HNF-3beta belongs to the winged-helix family of transcription factors and has been implicated in the control of cell-specific and developmental gene expression. The results of the present study show that the cell lineage-specific transcription factor HNF-3beta is an essential component of a novel protein kinase C response element in the glucagon gene.
Fürstenau
U
U
Department of Molecular Pharmacology, University of Göttingen, Germany.
Schwaninger
M
M
Blume
R
R
Kennerknecht
I
I
Knepel
W
W
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D002460:Cell Line; D002874:Chromosome Mapping; D000242:Cyclic AMP / Q000494:pharmacology; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005934:Glucagon / Q000235:genetics*; D051542:Hepatocyte Nuclear Factor 3-beta; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000235:genetics; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D018631:ras Proteins / Q000378:metabolism
D004268:DNA-Binding Proteins; C496503:Foxa2 protein, mouse; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D014157:Transcription Factors; D051542:Hepatocyte Nuclear Factor 3-beta; D005934:Glucagon; D000242:Cyclic AMP; D011493:Protein Kinase C; D018631:ras Proteins; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium