TY - JOUR
T1 - Glucagon-like peptide 1 stimulates insulin gene promoter activity by protein kinase A-independent activation of the rat insulin I gene cAMP response element
AU - Skoglund, Gunnar
AU - Hussain, Mehboob A.
AU - Holz, George G.
PY - 2000/7
Y1 - 2000/7
N2 - Glucagon-like peptide 1 (GLP-1), a hormonal activator of adenyl cyclase, stimulates insulin gene transcription, an effect mediated by the cAMP response element (CRE) of the rat insulin I gene promoter (RIP1). Here we demonstrate that the signaling mechanism underlying stimulatory effects of GLP-1 on insulin gene transcription results from protein kinase A (PKA)- independent activation of the RIP1 CRE. Although GLP-1 stimulates cAMP production in rat INS-1 insulinoma cells, we find accompanying activation of a -410-bp RIP1 luciferase construct (-410RIP1-LUC) to exist independently of tiffs second messenger. GLP-1 produced a dose-dependent stimulation of - 410RIP1-LUC (EC50 0.43 nmol/l), an effect reproduced by the GLP-1 receptor agonist exendin-4 and abolished by the antagonist exendin(9-39). Activation of RIP1 by GLP-1 was not affected by cotransfection with dominant-negative G(s) α, was not blocked by cAMP antagonist Rp-cAMPS, and was insensitive to PKA antagonist H-89. Truncation of -410RIP1-LUC to generate -307-, -206-, and -166-bp constructs revealed 2 segments of RIP1 targeted by GLP-1. The first segment, not regulated by forskolin, was located between -410 and -307 bp of the promoter. The second segment, regulated by both GLP-1 and forskolin, included the CRE and was located between -206 and -166 bp. Consistent with these observations, stimulatory effects of GLP-1 at RIP1 were reduced after introduction of Δ-182 and Δ-183/180 inactivating deletions at the CRE. The action of GLP-1 at -410RIP1-LUC was also reduced by cotransfection with A- CREB, a genetically engineered isoform of the CRE binding protein CREB, which dimerizes with and prevents binding of basic-region-leucine-zipper (bZIP) transcription factors to the CRE. In contrast, the action of GLP-1 at the CRE was not blocked by cotransfection with M1-CREB, an isoform that lacks a consensus serine residue serving as substrate for PKA-mediated phosphorylation. On the basis of these studies, it is proposed that PKA- independent stimulatory actions of GLP-1 at RIP1 are mediated by bZIP transcription factors related in structure but not identical to CREB.
AB - Glucagon-like peptide 1 (GLP-1), a hormonal activator of adenyl cyclase, stimulates insulin gene transcription, an effect mediated by the cAMP response element (CRE) of the rat insulin I gene promoter (RIP1). Here we demonstrate that the signaling mechanism underlying stimulatory effects of GLP-1 on insulin gene transcription results from protein kinase A (PKA)- independent activation of the RIP1 CRE. Although GLP-1 stimulates cAMP production in rat INS-1 insulinoma cells, we find accompanying activation of a -410-bp RIP1 luciferase construct (-410RIP1-LUC) to exist independently of tiffs second messenger. GLP-1 produced a dose-dependent stimulation of - 410RIP1-LUC (EC50 0.43 nmol/l), an effect reproduced by the GLP-1 receptor agonist exendin-4 and abolished by the antagonist exendin(9-39). Activation of RIP1 by GLP-1 was not affected by cotransfection with dominant-negative G(s) α, was not blocked by cAMP antagonist Rp-cAMPS, and was insensitive to PKA antagonist H-89. Truncation of -410RIP1-LUC to generate -307-, -206-, and -166-bp constructs revealed 2 segments of RIP1 targeted by GLP-1. The first segment, not regulated by forskolin, was located between -410 and -307 bp of the promoter. The second segment, regulated by both GLP-1 and forskolin, included the CRE and was located between -206 and -166 bp. Consistent with these observations, stimulatory effects of GLP-1 at RIP1 were reduced after introduction of Δ-182 and Δ-183/180 inactivating deletions at the CRE. The action of GLP-1 at -410RIP1-LUC was also reduced by cotransfection with A- CREB, a genetically engineered isoform of the CRE binding protein CREB, which dimerizes with and prevents binding of basic-region-leucine-zipper (bZIP) transcription factors to the CRE. In contrast, the action of GLP-1 at the CRE was not blocked by cotransfection with M1-CREB, an isoform that lacks a consensus serine residue serving as substrate for PKA-mediated phosphorylation. On the basis of these studies, it is proposed that PKA- independent stimulatory actions of GLP-1 at RIP1 are mediated by bZIP transcription factors related in structure but not identical to CREB.
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U2 - 10.2337/diabetes.49.7.1156
DO - 10.2337/diabetes.49.7.1156
M3 - Article
C2 - 10909973
AN - SCOPUS:0033947157
SN - 0012-1797
VL - 49
SP - 1156
EP - 1164
JO - Diabetes
JF - Diabetes
IS - 7
ER -