Proximal cyclic AMP response element is essential for exendin-4 induction of rat EGR-1 gene.
Glucagon-like peptide-1 and its potent agonist exendin-4 induce several immediate early response genes (IEGs) that code for transcription factors implicated in cell proliferation, differentiation, and apoptosis. We recently observed that early growth response factor-1 (EGR-1), an IEG product, was required for transcriptional activation of Ccnd1 (cyclin D1) gene by exendin-4. Herein, the regulatory mechanism whereby exendin-4 activates the transcription of EGR-1 gene was investigated in the pancreatic beta-cell line INS-1. Deletion analysis of rat EGR-1 promoter identified a critical region between -73 and -46 for the activation of EGR-1 in response to exendin-4. Mutation of the proximal putative cAMP response element (CRE, 5'-GTACGTCA-3') located at -69 resulted in a significant decrease in the EGR-1 transcription, whereas the mutation of the distal putative CRE at -139 was without such an effect. In immune supershift assays using exendin-4-treated cells, binding of cAMP response element-binding protein (CREB) phosphorylated on Ser(133) to the proximal CRE was increased. Employment of a CREB mutant containing Ala substitution at Ser(133) or a dominant negative CREB mutant that inhibits the binding of endogenous CREB to DNA significantly decreased the exendin-4-induced EGR-1 transcription. In experiments using specific protein kinase inhibitors, the effect of H-89 was more prominent than PD-98059, indicating the predominance of the PKA signaling over the MEK/ERK in induction of EGR-1. Therefore, it appears that the proximal CRE site is critical and the binding with CREB phosphorylated on Ser(133) is necessary for induction of the EGR-1 transcription by exendin-4.
Kang
Jung-Hoon
JH
Dept. of Physiology, The Catholic University of Korea, Seoul 137-701, Korea. yhjo@catholic.ac.kr
Kim
Myung-Jun
MJ
Jang
Hwa-In
HI
Koh
Kyung-Hee
KH
Yum
Keun-Sang
KS
Rhie
Duck-Joo
DJ
Yoon
Shin Hee
SH
Hahn
Sang June
SJ
Kim
Myung-Suk
MS
Jo
Yang-Hyeok
YH
D000818:Animals; D002478:Cells, Cultured; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D051766:Early Growth Response Protein 1 / Q000235:genetics* / Q000378:metabolism; D000077270:Exenatide; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D009154:Mutation; D010455:Peptides / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects*; D014688:Venoms / Q000494:pharmacology*
D017362:Cyclic AMP Response Element-Binding Protein; D051766:Early Growth Response Protein 1; C486110:Egr1 protein, rat; D010455:Peptides; D014688:Venoms; D000077270:Exenatide; D017868:Cyclic AMP-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases