Exendin-4 as a stimulator of rat insulin I gene promoter activity via bZIP/CRE interactions sensitive to serine/threonine protein kinase inhibitor Ro 31-8220

Endocrinology. 2002 Jun;143(6):2303-13. doi: 10.1210/endo.143.6.8870.


Signal transduction properties of exendin-4 (Ex-4) underlying its ability to stimulate rat insulin I gene promoter (RIP1) activity were assessed in the pancreatic beta-cell line INS-1. Ex-4 acted via glucagon-like peptide-1 receptors to stimulate RIP1 in a glucose-dependent manner, as measured in cells transfected with a -410-bp RIP1-luciferase construct (RIP1-Luc). The action of Ex-4 was independent of cAMP and PKA because it was not blocked by cotransfection with dominant-negative G alpha(s), was unaffected by pretreatment with the membrane-permeant cAMP antagonist 8-Br-Rp-cAMPS, and remained apparent after treatment with PKA inhibitors H-89 or KT 5720. Similarly, cotransfection with a dominant-negative isoform of the type-2 cAMP-regulated guanine nucleotide exchange factor (Epac2) failed to alter the response to Ex-4. Ro 31-8220, a serine/threonine protein kinase inhibitor that targets PKC as as well as the 90-kDa ribosomal S6 kinase (RSK) and mitogen- and stress-activated protein kinase (MSK) family of cAMP response element-binding protein (CREB) kinases, blocked the stimulatory action of Ex-4 at RIP1-Luc. However, selective inhibition of PKC using K-252c, prolonged exposure to phorbol 1,2-myristate-13-acetate, or cotransfection with dominant-negative atypical PKC-zeta, was without effect. A-CREB, a dominant-negative inhibitor of basic region-leucine zipper transcription factors (bZIPs) related in structure to CREB, inhibited the action of Ex-4 at RIP1-Luc, whereas A-ATF-2 was ineffective. Similarly, introduction of deletions at the RIP1 cAMP response element (CRE), or truncation of RIP1 to remove the CRE, nearly abolished the action of Ex-4. Inactivating mutations introduced at the A4/A3 elements, binding sites for the glucose-regulated homeodomain transcription factor PDX-1, did not diminish the response to Ex-4, although a marked reduction of basal promoter activity was observed. The glucose-dependent stimulation of RIP1-Luc by Ex-4 was reproduced using a synthetic reporter (RIP1-CRE-Luc) incorporating multimerized CREs of the RIP1 nonpalindromic sequence 5'-TGACGTCC-3'. It is concluded that the bZIP and CRE-mediated stimulation of RIP1 by Ex-4 explains, at least in part, how this insulinotropic hormone facilitates transcriptional activity of the rat insulin I gene.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Basic-Leucine Zipper Transcription Factors
  • Cells, Cultured
  • Cyclic AMP / physiology
  • Cyclic AMP Response Element-Binding Protein / physiology
  • Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
  • DNA-Binding Proteins / genetics*
  • Enzyme Inhibitors / pharmacology*
  • Exenatide
  • G-Box Binding Factors
  • Glucose / physiology
  • Humans
  • Indicators and Reagents
  • Indoles / pharmacology*
  • Insulin / genetics
  • Insulin-Like Growth Factor I / genetics*
  • Luciferases
  • Peptides / pharmacology*
  • Plasmids / genetics
  • Promoter Regions, Genetic / genetics
  • Protein Serine-Threonine Kinases / antagonists & inhibitors*
  • Rats
  • Signal Transduction / drug effects
  • Stimulation, Chemical
  • Transcription Factors / genetics*
  • Transfection
  • Venoms*


  • Basic-Leucine Zipper Transcription Factors
  • Cyclic AMP Response Element-Binding Protein
  • DNA-Binding Proteins
  • Enzyme Inhibitors
  • G-Box Binding Factors
  • Indicators and Reagents
  • Indoles
  • Insulin
  • Peptides
  • Transcription Factors
  • Venoms
  • Insulin-Like Growth Factor I
  • Exenatide
  • Cyclic AMP
  • Luciferases
  • Protein Serine-Threonine Kinases
  • Cyclic AMP-Dependent Protein Kinases
  • Glucose
  • Ro 31-8220