Group VIA phospholipase A2 forms a signaling complex with the calcium/calmodulin-dependent protein kinase IIbeta expressed in pancreatic islet beta-cells.

Abstract

Insulin-secreting pancreatic islet beta-cells express a Group VIA Ca(2+)-independent phospholipase A(2) (iPLA(2)beta) that contains a calmodulin binding site and protein interaction domains. We identified Ca(2+)/calmodulin-dependent protein kinase IIbeta (CaMKIIbeta) as a potential iPLA(2)beta-interacting protein by yeast two-hybrid screening of a cDNA library using iPLA(2)beta cDNA as bait. Cloning CaMKIIbeta cDNA from a rat islet library revealed that one dominant CaMKIIbeta isoform mRNA is expressed by adult islets and is not observed in brain or neonatal islets and that there is high conservation of the isoform expressed by rat and human beta-cells. Binary two-hybrid assays using DNA encoding this isoform as bait and iPLA(2)beta DNA as prey confirmed interaction of the enzymes, as did assays with CaMKIIbeta as prey and iPLA(2)beta bait. His-tagged CaMKIIbeta immobilized on metal affinity matrices bound iPLA(2)beta, and this did not require exogenous calmodulin and was not prevented by a calmodulin antagonist or the Ca(2+) chelator EGTA. Activities of both enzymes increased upon their association, and iPLA(2)beta reaction products reduced CaMKIIbeta activity. Both the iPLA(2)beta inhibitor bromoenol lactone and the CaMKIIbeta inhibitor KN93 reduced arachidonate release from INS-1 insulinoma cells, and both inhibit insulin secretion. CaMKIIbeta and iPLA(2)beta can be coimmunoprecipitated from INS-1 cells, and forskolin, which amplifies glucose-induced insulin secretion, increases the abundance of the immunoprecipitatable complex. These findings suggest that iPLA(2)beta and CaMKIIbeta form a signaling complex in beta-cells, consistent with reports that both enzymes participate in insulin secretion and that their expression is coinduced upon differentiation of pancreatic progenitor to endocrine progenitor cells.

• Authors: Zhepeng Wang, Sasanka Ramanadham, Zhongmin Alex Ma, Shunzhong Bao, David J Mancuso, Richard W Gross, John Turk
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 280 Issue 8 Pg. 6840-9 (Feb 25 2005) ISSN: 0021-9258 [Print] United States
• PMID: 15576376 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Calmodulin
• Enzyme Inhibitors
• Insulin
• RNA, Messenger
• Arachidonic Acid
• CAMK2B protein, human
• Calcium-Calmodulin-Dependent Protein Kinase Type 2
• Calcium-Calmodulin-Dependent Protein Kinases
• Camk2b protein, rat
• Phospholipases A
• Group VI Phospholipases A2
• PLA2G6 protein, human
• Phospholipases A2

Topics

• Animals
• Arachidonic Acid (metabolism)
• Calcium-Calmodulin-Dependent Protein Kinase Type 2
• Calcium-Calmodulin-Dependent Protein Kinases (metabolism, physiology)
• Calmodulin (pharmacology)
• Cell Line
• Enzyme Inhibitors (pharmacology)
• Group VI Phospholipases A2
• Insecta
• Insulin (metabolism)
• Insulin Secretion
• Islets of Langerhans (enzymology, metabolism)
• Phospholipases A (metabolism, physiology)
• Phospholipases A2
• Protein Binding
• RNA, Messenger (analysis)
• Rats
• Signal Transduction
• Tissue Distribution
• Two-Hybrid System Techniques