Regulation by glucagon of the rat histidase gene promoter in cultured rat hepatocytes and human hepatoblastoma cells.
Histidase (Hal), the amino acid-degrading enzyme of histidine, is regulated by the protein content of the diet and by hormones such as glucocorticoids and glucagon. However, glucagon can activate the following two possible transduction pathways: protein kinase A (PKA) and protein kinase C (PKC). The aim of this study was to isolate the 5'-flanking region of rat Hal gene to locate possible cAMP- and glucocorticoid-responsive elements and to identify whether the activation of the Hal promoter by glucagon occurs via PKA or PKC. The results showed that glucagon was able to induce Hal expression 1.5-fold in primary hepatocytes. The addition of phorbol 12-myristate,13-acetate (PMA) and forskolin to hepatocytes increased Hal mRNA concentration by 100 and 40%, respectively. To identify the Hal gene regulatory region, a 1248-bp fragment of the 5'-region was obtained. The transcription initiation site was located at 404 bp from ATG. The sequence did not show consensus TATA-like or CAAT-like boxes in the first 100 bp upstream from the transcription start site. The promoter contained six GC rich boxes, seven putative AP1 binding sites, and four glucocorticoid-responsive elements. The putative Hal promoter region was cloned into the pGL3basic vector and transfected into HepG2 cells. Luciferase expression was significantly stimulated by glucagon (0.9-fold), forskolin (0.9-fold), PMA (2.0-fold), and dexamethasone (2.9-fold). This evidence supports that the Hal gene is turned on by glucocorticoids and by glucagon either via PKC or PKA, but prefers the PKA pathway.
Alemán
Gabriela
G
Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Departamento Fisiología de la Nutrición, Universidad Nacional Autonomica de México, Mexico City, México.
Ortíz
Victor
V
Langley
Elizabeth
E
Tovar
Armando R
AR
Torres
Nimbe
N
D000818:Animals; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects; D005934:Glucagon / Q000008:administration & dosage*; D018197:Hepatoblastoma / Q000201:enzymology*; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology*; D006638:Histidine Ammonia-Lyase / Q000378:metabolism*; D006801:Humans; D008297:Male; D011401:Promoter Regions, Genetic / Q000235:genetics; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction / Q000187:drug effects
D005934:Glucagon; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D006638:Histidine Ammonia-Lyase