Interleukin-1P induces gene expression and secretion of group-I1 phospholipase A, and release of prostaglandin E2 from rat mesangial cells. The interleukin-lp-induced synthesis of group-I1 phospholipase A2 is prevented by transforming growth factor-/?,, whereas transforming growth factor-p2 potentiated the interleukin-lp-evoked prostaglandin E, production. Transforming growth factor-p, itself did not induce synthesis of group-I1 phospholipase A,, although it stimulated prostaglandin E2 formation. Here we describe the effect of interleukin-lp and transforming growth factorp2 on a cytosolic phospholipase A2 activity and prostaglandin E2 formation in rat mesangial cells.Based on the resistance to dithiothreitol and migration profiles on a Mono-Q anion-exchange column and a Superose 12 gel-filtration column, the cytosolic phospholipase A2 activity was assigned to a high-molecular-mass phospholipase A,. Measured with l-stearoyl-2-[1 -'4C]arachidonoylglycerophosphocholine as substrate, both interleukin-1p and transforming growth factor-pz enhanced the high-molecular-mass phospholipase A, activity. The stimulation of rat mesangial cells with interleukin-1 p and transforming growth factor-pz was time-and dose-dependent with maximal cytosolic phospholipase A, activities at 10 nM and at 10 ng/ml respectively, after 24 h of stimulation. Under these conditions, interleukin-1 p and transforming growth factor-p2 enhanced the cytosolic phospholipase A, activity 2.2 f 0.6-fold and 2.5 f 0.6-fold, respectively. These results strongly suggest that an enhanced cytosolic high-molecular-mass phospholipase A, activity is involved in the formation of prostaglandin E, mediated by transforming growth factor-p2. Whether interleukin-lp induced group-I1 phospholipase A, and/or interleukin-lp-enhanced cytosolic phospholipase A, activity is involved in prostaglandin E2 formation in rat mesangial cells is discussed.Phospholipase A, (PLA2) are a diverse family of important enzymes which have attracted considerable attention because of their role in the production of potent inflammatory mediators such as prostaglandins, leukotrienes and plateletactivating factor [ 11. Mammalian cells contain several PLA, enzymes including the 14-kDa enzymes and the more recently described high-molecular-mass enzymes. The 14-kDa PLA2s can be discriminated into group I and group I1 based on their primary structure [2]. Despite the potential importance of the PLA2s in lipid inflammatory mediator production, little is known about the type of PLA, involved.We and others have previously shown that the pro-inflammatory cytokines interleukin-1P (IL-1p) and tumor necrosis factor stimulate the synthesis and secretion of group-I1 PLA, Correspondence to H. van

CAPRI is a member of the GAP1 family of GTPase-activating proteins (GAPs) for small G proteins. It is known to function as an amplitude sensor for intracellular Ca 2؉ levels stimulated by extracellular signals and has a catalytic domain with dual Ras-GAP and RapGAP activities. Here, we have investigated the mechanism that switches CAPRI between its two GAP activities. We demonstrate that CAPRI forms homodimers in vitro and in vivo in a Ca 2؉ -dependent manner. The site required for dimerization was pinpointed by deletion and point mutations to a helix motif that forms a hydrophobic face in the extreme C-terminal tail of the CAPRI protein. Deletion of this helix motif abolished dimer formation but did not affect translocation of CAPRI to the plasma membrane upon cell stimulation with histamine. We found that dimeric and monomeric CAPRI coexist in cells and that the ratio of dimeric to monomeric CAPRI increases upon cell stimulation with histamine. Free Ca 2؉ at physiologically relevant concentrations was both necessary and sufficient for dimer formation. Importantly, the monomeric and dimeric forms of CAPRI exhibited differential GAP activities in vivo; the wild-type form of CAPRI had stronger RapGAP activity than RasGAP activity, whereas a monomeric CAPRI mutant showed stronger RasGAP than RapGAP activity. These results demonstrate that CAPRI switches between its dual GAP roles by forming monomers or homodimers through a process regulated by Ca 2؉ . We propose that Ca 2؉ -dependent dimerization of CAPRI may serve to coordinate Ras and Rap1 signaling pathways.The closely related small G proteins (GTPases) Ras and Rap1 are conserved molecular switches that couple extracellular signals to a wide range of cellular responses through different signaling networks (1). Ras plays a central role in cell proliferation and cell survival and is a major oncogene (2). Rap1 was originally identified as a protein that reverts the effects of active Ras, such as the loss of adhesion accompanying cell transformation by oncogenic K-Ras (3) or the activation of ERK and ELK1 (4, 5). However, subsequent studies showed that Rap1 is not a mere anti-Ras protein but has discrete functions, notably in integrinmediated cell adhesion and spreading, formation of cell/cell contacts (6), superoxide formation, and cAMP-induced neurite outgrowth (1, 7). These distinct physiological roles of Ras and Rap1 are mediated by their differential use of effector proteins and also through differential subcellular localizations of their effectors (8).As is typical for small G proteins, the biological activities of both Ras and Rap1 are controlled by a GDP/GTP cycle; they are active in their GTP-bound form and inactive in their GDPbound form (1). Guanine nucleotide exchange factors activate them by promoting the dissociation of GDP, allowing excess free cellular GTP to bind, whereas GAPs 2 inactivate them by stimulating their intrinsic GTPase activity. A number of guanine nucleotide exchange factors and GAPs for Ras and Rap have been identified. In general, t...