title: Calcium/calmodulin-dependent kinase IV controls glucose-induced Irs2 expression in mouse beta cells via activation of cAMP response element-binding protein.
abstract: OBJECTIVE
  Irs2, which is upregulated by glucose, is important for beta cell plasticity. Cyclic AMP response element-binding protein (CREB) stimulates beta cell Irs2 expression and is a major calcium/calmodulin-dependent kinase (CaMK)(IV) target in neurons. We therefore hypothesised that CaMK(IV) mediates glucose-induced Irs2 expression in beta cells via CREB activation.
  
  
  METHODS
  The functions of CaMK(IV) and CREB were investigated in MIN6 beta cells and mouse islets using the CaMK inhibitor KN62, the calcium chelator bapta-(AM) and the voltage-dependent calcium channel inhibitor nifedipine. Small interfering RNAs were used to silence endogenous CaMK(IV) production and expression vectors to overproduce constitutively active and dominant negative forms of CaMK(IV) and CREB. Irs1 and Irs2 expression were determined by quantitative PCR and Western blotting, and the role of CREB was also investigated by assessing its phosphorylation on serine 133.
  
  
  RESULTS
  Increasing the glucose concentration from 2.5 to 25 mmol/l stimulated CREB phosphorylation on serine 133 and specifically stimulated Irs2 but not Irs1 expression. Similarly, overproduction of a constitutively active form of CaMK(IV) promoted sustained CREB phosphorylation and a significant increase in Irs2 but not Irs1 expression. In contrast, these stimulatory effects of glucose were all suppressed by overproducing an inactive CaMK(IV) mutant. Inhibition of glucose-induced calcium influx with nifedipine or chelation of intracellular calcium with bapta-(AM), as well as silencing of CaMK(IV) or inhibition of its activity with KN62 resulted in similar observations. Finally, overproduction of a dominant negative form of CREB completely suppressed glucose and CaMK(IV) stimulation of Irs2 expression.
  
  
  CONCLUSIONS
  Our results suggest that the Ca(2+)/CaMK(IV)/CREB cascade plays a critical role in the regulation of Irs2 expression in beta cells.
authors.affiliation: Diabetes Research Group, School of Medicine, King's College London, Guy's Campus, London, SE1 1UL, UK.
authors.forename: S J
authors.initials: SJ
authors.lastname: Persaud
authors.forename: B
authors.initials: B
authors.lastname: Liu
authors.forename: H Barbosa
authors.initials: HB
authors.lastname: Sampaio
authors.forename: P M
authors.initials: PM
authors.lastname: Jones
authors.forename: D S
authors.initials: DS
authors.lastname: Muller
mesh_terms: D000818:Animals; D015153:Blotting, Western; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4 / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000378:metabolism*; D005947:Glucose / Q000494:pharmacology*; D066298:In Vitro Techniques; D055504:Insulin Receptor Substrate Proteins / Q000235:genetics / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000187:drug effects* / Q000378:metabolism*; D051379:Mice; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D055504:Insulin Receptor Substrate Proteins; C526298:Irs2 protein, mouse; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D005947:Glucose