Vol. 298, Issue 1, 272-278, July 2001

Ca²⁺/Calmodulin-Dependent Protein Kinase II and Cytosolic Phospholipase A₂ Contribute to Mitogenic Signaling in Myeloblastic Leukemia U-937 Cells

Mubarack M. Muthalif, Farid Ljuca, J. Brent Roaten, Neelu Pentapaty, Mohammed R. Uddin and Kafait U. Malik

Department of Pharmacology, College of Medicine, The University of Tennessee, Memphis, Tennessee

The signaling mechanisms downstream of growth factor-stimulated proliferation in myeloid leukemia cells have not yet been fully elucidated. Recent evidence suggests that alternate pathways to the mitogen-activated protein kinase cascade are required. We have previously shown that Ca²⁺/calmodulin-dependent protein kinase II (CaM kinase II) activates cytosolic phospholipase A₂ (cPLA₂), which is involved in the proliferation of vascular smooth muscle cells. In the present study, the contribution of this pathway was investigated in the proliferation of U-937 myeloid leukemia cells. In U-937 cells, fetal bovine serum (FBS)-induced proliferation was attenuated by CaM kinase II inhibitor KN-93 but not by its inactive analog KN-92. Inhibitors of cPLA₂ (methyl arachidonyl fluorophosphonate and arachidonyl trifluoromethyl ketone) also reduced proliferation of U-937 cells. FBS-induced proliferation was also attenuated by cotransfection with cPLA₂ antisense oligonucleotides. These results suggest a role for CaM kinase II and cPLA₂ in the proliferation of U-937 cells. FBS stimulated CaM kinase II and cPLA₂ activities in a time-dependent manner. Moreover, FBS-stimulated phosphorylation and activation of cPLA₂ activation was inhibited by KN-93. FBS-stimulated phosphorylation of CaM kinase II was blocked by KN-93 but not by cPLA₂ inhibitors, suggesting that CaM kinase II activates cPLA₂. The products of phospholipid hydrolysis produced by cPLA₂, lysophosphatidylcholine but not arachidonic acid, increased [³H]thymidine incorporation in U-937 cells. These data suggest that exposure of U-937 cells to FBS promotes phosphorylation and activation of CaM kinase II, leading to stimulation of cPLA₂ and generation of lysophosphatidylcholine and resultant proliferation of these cells.