No abstract

Cytosolic phospholipase A(2) (cPLA(2)) is of special interest because it selectively releases arachidonic acid from membrane phospholipids. Arachidonic acid has been implicated to play an important role in various cellular responses. Recently arachidonic acid release and prostaglandin synthesis have been shown to be cell cycle dependent and therefore the activity of cPLA(2) during the ongoing cell cycle was investigated, using the mitotic shake off method for cell synchronisation. cPLA(2) activity was high in mitotic cells and decreased rapidly in the early G1 phase. A strong increase in activity was measured following the G1/S transition in both neuroblastoma and Chinese hamster ovary cells. The changes in activity were not due to a difference in cPLA(2) expression but due to phosphorylation of cPLA(2). Phosphorylation of cPLA(2) occurs through MAPK since the use of a specific MAPK kinase inhibitor and serum depletion of synchronised cells inhibited cPLA(2) activity.

Arachidonic acid has been implicated to play a role in physiological and pathophysiological processes and is selectively released by the 85-kDa cytosolic phospholipase A 2 (cPLA 2 ).

Arachidonic acid has been implicated in regulating cellular proliferation, and is preferentially released by the 85-kDa cytosolic phospholipase A2 (cPLA2). Recently, we demonstrated that cPLA2 is activated at distinct periods during the ongoing cell cycle of neuroblastoma cells. The purpose of the present study was to establish the role of these cPLA2 activity peaks in cell cycle progression. Inhibition of cPLA2 activity with arachidonyl trifluoromethylketone (ATK) in early G1 phase reduced DNA synthesis markedly. A 24-h incubation with ATK revealed no significant difference in cell number compared to untreated cells, although cPLA2 activity was still inhibited. This suggests redundancy of different PLA2 enzymes. Lipoxygenase inhibition in early G1 resulted in G1 phase arrest, whereas inhibitors for cyclooxygenase had no effect. Furthermore, cells stopped progressing through S phase when lipoxygenase was inhibited in early S phase, demonstrating the requirement of lipoxygenase products for S phase progression.

cPLA 2 plays a key role in many signal transduction cascades by hydrolyzing arachidonic acid from membrane phospholipids. Tight control of cPLA 2 activity by a number of regulatory mechanisms is essential to its cellular function. We recently described the localization of cPLA 2 in clusters in fibroblasts and now propose that these clusters reflect a localized inactive pool from which active monomers can be recruited to keep cPLA 2 activity under control on the subcellular level. Using an electron microscopic in vitro approach, we show that cPLA 2 monomers, but not the clusters, bind to membranes in a Ca 2+dependent manner. This binding is accompanied by hydrolytic activity. The present data combined with our previous observation of a relative abundance of clusters over monomers in fixed fibroblasts [Bunt,