title: Protein hydrolysates stimulate proglucagon gene transcription in intestinal endocrine cells via two elements related to cyclic AMP response element.
abstract: OBJECTIVE
  Protein hydrolysates (peptones) increase not only glucagon-like peptide-1 (GLP-1) secretion but also transcription of the proglucagon ( PG) gene in the intestine. The critical physiological roles of gut-derived GLPs raised hope for their therapeutic use in several disorders, especially GLP-1 in diabetes. We aimed to investigate the molecular mechanisms involved in this nutrient- PG gene interaction.
  
  
  METHODS
  Wild-type and mutated PG promoter fragments fused to the luciferase reporter gene were transfected into enteroendocrine STC-1 cells, which were then either treated or not with peptones. Co-transfection with expression vectors of dominant-negative forms of cAMP response element binding protein (CREB) and protein kinase A (PKA) proteins were performed, as well as electrophoresis mobility shift assays.
  
  
  RESULTS
  Deletion analysis showed that the promoter region spanning between -350 and -292 bp was crucial for the transcriptional stimulation induced by peptones. Site-directed mutagenesis of the canonical cAMP response element (CRE(PG)) and of the adjacent putative CRE site (CRE-like1) led to a dramatic inhibition of the promoter responsiveness to peptones. Over expression of a dominant-negative mutant of CREB or of PKA produced a comparable and selective inhibitory effect on the activity of transfected promoter fragment containing the -350/-292 sequence. EMSA showed that CREB and fra2 transcription factors bound to CRE(PG) and CRE-like1 elements respectively, independently of peptone treatment.
  
  
  CONCLUSIONS
  Our report identified cis- and trans-regulatory elements implicated in the transcriptional control of PG gene by nutrients in enteroendocrine cells. It highlights the role of a previously unsuspected CRE-like1 element, and emphasises the importance of CRE-related sequences in the regulation of PG gene transcription in the intestine.
authors.affiliation: INSERM Unité 45, IFR 62, Faculté de Médecine RTH Laënnec, Lyon, France.
authors.forename: J-C
authors.initials: JC
authors.lastname: Gevrey
authors.forename: M
authors.initials: M
authors.lastname: Malapel
authors.forename: J
authors.initials: J
authors.lastname: Philippe
authors.forename: G
authors.initials: G
authors.lastname: Mithieux
authors.forename: J-A
authors.initials: JA
authors.lastname: Chayvialle
authors.forename: J
authors.initials: J
authors.lastname: Abello
authors.forename: M
authors.initials: M
authors.lastname: Cordier-Bussat
mesh_terms: D000818:Animals; D001483:Base Sequence; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D003001:Cloning, Molecular; D017362:Cyclic AMP Response Element-Binding Protein / Q000502:physiology*; D017931:DNA Primers; D005934:Glucagon / Q000235:genetics*; D007422:Intestines; D051379:Mice; D008822:Mice, Transgenic; D016297:Mutagenesis, Site-Directed; D052336:Proglucagon; D011401:Promoter Regions, Genetic; D011498:Protein Precursors / Q000235:genetics*; D017384:Sequence Deletion; D014158:Transcription, Genetic
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D017931:DNA Primers; D011498:Protein Precursors; D052336:Proglucagon; D005934:Glucagon