title: Glucose-dependent downregulation of glucagon gene expression mediated by selective interactions between ALX3 and PAX6 in mouse alpha cells.
abstract: OBJECTIVE
  The stimulation of glucagon secretion in response to decreased glucose levels has been studied extensively. In contrast, little is known about the regulation of glucagon gene expression in response to fluctuations in glucose concentration. Paired box 6 (PAX6) is a key transcription factor that regulates the glucagon promoter by binding to the G1 and G3 elements. Here, we investigated the role of the transcription factor aristaless-like homeobox 3 (ALX3) as a glucose-dependent modulator of PAX6 activity in alpha cells.
  
  
  METHODS
  Experiments were performed in wild-type or Alx3-deficient islets and alphaTC1 cells. We used chromatin immunoprecipitations and electrophoretic mobility shift assays for DNA binding, immunoprecipitations and pull-down assays for protein interactions, transfected cells for promoter activity, and small interfering RNA and quantitative RT-PCR for gene expression.
  
  
  RESULTS
  Elevated glucose concentration resulted in stimulated expression of Alx3 and decreased glucagon gene expression in wild-type islets. In ALX3-deficient islets, basal glucagon levels were non-responsive to changes in glucose concentration. In basal conditions ALX3 bound to the glucagon promoter at G3, but not at G1. ALX3 could form heterodimers with PAX6 that were permissive for binding to G3 but not to G1. Thus, increasing the levels of ALX3 in response to glucose resulted in the sequestration of PAX6 by ALX3 for binding to G1, thus reducing glucagon promoter activation and glucagon gene expression.
  
  
  CONCLUSIONS
  Glucose-stimulated expression of ALX3 in alpha cells provides a regulatory mechanism for the downregulation of glucagon gene expression by interfering with PAX6-mediated transactivation on the glucagon G1 promoter element.
authors.affiliation: Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Madrid, Spain.|Instituto de Investigaciones Biomédicas Alberto Sols, Consejo Superior de Investigaciones Científicas/Universidad Autónoma de Madrid (CSIC/UAM), Calle Arturo Duperier 4, 28029, Madrid, Spain.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Mirasierra
authors.affiliation: Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Madrid, Spain. mvallejo@iib.uam.es.|Instituto de Investigaciones Biomédicas Alberto Sols, Consejo Superior de Investigaciones Científicas/Universidad Autónoma de Madrid (CSIC/UAM), Calle Arturo Duperier 4, 28029, Madrid, Spain. mvallejo@iib.uam.es.
authors.forename: Mario
authors.initials: M
authors.lastname: Vallejo
mesh_terms: D000818:Animals; D002460:Cell Line; D024202:Electrophoretic Mobility Shift Assay; D005934:Glucagon / Q000378:metabolism*; D050416:Glucagon-Secreting Cells / Q000378:metabolism*; D005947:Glucose / Q000494:pharmacology*; D018398:Homeodomain Proteins / Q000378:metabolism*; D047468:Immunoprecipitation; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D000071841:PAX6 Transcription Factor / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: C113607:Alx3 protein, mouse; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; C000605732:Pax6 protein, mouse; D005934:Glucagon; D005947:Glucose
keywords: Alpha cells; Glucagon; Glucose-dependent gene expression; Homeodomain; Paired domain; Transcription