CLONING AND EXPRESSION OF A GROUP-IV CYTOSOLIC CA2-DEPENDENT PHOSPHOLIPASE-A(2) FROM RAT PANCREATIC-ISLETS - COMPARISON OF THE EXPRESSED ACTIVITY WITH THAT OF AN ISLET GROUP-VI CYTOSOLIC CA2+-INDEPENDENT PHOSPHOLIPASE-A(2)()

Stimulation of pancreatic islets with glucose induces phospholipid hyd rolysis and accumulation of nonesterified arachidonic acid, which may play signaling or effector roles in insulin secretion. Of enzymes that catalyze phospholipid hydrolysis, islet beta-cells express low molecu lar weight secretory phospholipases A(2) (PLA(2)) and a Group VI, Ca2-independent PLA(2)(iPLA(2)). Previous studies indicate that islets al so express a protein recognized by antibodies against a Group IV, cyto solic, Ca2+-dependent PLA(2) (cPLA(2)), To further examine the possibl e expression of cPLA(2) by islets, we screened a rat islet cDNA librar y with a probe that recognizes cPLA(2) sequence, and isolated a full-l ength cPLA(2) cDNA. The rat islet cPLA(2)-deduced amino acid sequence is 96% identical to those of human and mouse cPLA(2). Transfection of COS-7 cells with cPLA(2) cDNA in an expression vector induced expressi on of Ca2+-dependent PLA(2) activity and of a protein recognized by an ti-cPLA(2) antibody. Comparison of recombinant islet cPLA(2) and iPLA( 2) activities expressed in transfected COS-7 cells indicated that iPLA (2) but not cPLA(2) is stimulated by ATP. Both activities are similarl y sensitive to inhibition by arachidonyltrifluoromethyl ketone, but iP LA(2) is more effectively inhibited by a haloenol lactone suicide subs trate than cPLA(2). RT-PCR experiments with RNA from purified islet be ta-cells and from an alpha-cell-enriched population prepared by fluore scence-activated cell-sorting indicated that cPLA(2) mRNA is more abun dant in the beta-cell population. Immunoblotting analyses indicate tha t islets express cPLA(2)-immunoreactive protein, and that interleukin- 1 does not affect its expression.