Redundant and segregated functions of granule-associated heparin-binding group II subfamily of secretory phospholipases A(2) in the regulation of degranulation and prostaglandin D-2 synthesis in mast cells

We herein demonstrate that mast cells express all known members of the grou p II subfamily of secretory phospholipase A, (sPLA(2)) isozymes, and those having heparin affinity markedly enhance the exocytotic response. Rat masto cytoma RBL-2H3 cells transfected with heparin-binding (sPLA(2)-IIA, -V, and -IID), but not heparin-nonbinding (sPLA(2)-IIC), enzymes released more gra nule-associated markers (beta-hexosaminidase and histamine) than mock- or c ytosolic PLA(2)alpha (cPLA(2)alpha)-transfected cells after stimulation wit h IgE and Ag. Site-directed mutagenesis of sPLA(2)-IIA and -V revealed that both the catalytic and heparin-binding domains are essential for this func tion. Confocal laser and electron microscopic analyses revealed that sPLA(2 )-IIA, which was stored in secretory granules in unstimulated cells, accumu lated on the membranous sites where fusion between the plasma membrane and granule membranes occurred in activated cells. These results suggest that t he heparin-binding sPLA(2)s bind to the perigranular membranes through thei r heparin-binding domain, and lysophospholipids produced in situ by their e nzymatic action may facilitate the ongoing membrane fusion, In contrast to the redundant role of sPLA(2)-IIA, -IID, and -V in the regulation of degran ulation, only sPLA(2)-V had the ability to markedly augment IgE/Ag-stimulat ed immediate PGD(2) production, which reached a level comparable to that el icited by cPLA(2)alpha. The latter observation reveals an unexplored functi onal segregation among the three related isozymes expressed in the same cel l population.