The Role of Calcium and Phosphorylation of Cytosolic Phospholipase A2 in Regulating Arachidonic Acid Release in Macrophages

Abstract

Arachidonic acid release is induced in macrophages with diverse agonists including calcium ionophores, phorbol myristate acetate (PMA), okadaic acid, and the phagocytic particle, zymosan, and correlates with activation of cytosolic phospholipase A 2 (cPLA 2 ). The role of calcium and phosphorylation of cPLA 2 in regulating arachidonic acid release was investigated. Zymosan induced a rapid and transient increase in [Ca 2؉ ] i . This in itself is not sufficient to induce arachidonic acid release since ATP and platelet activating factor (PAF), agonists that induce transient calcium mobilization in macrophages, induced little arachidonic acid release. Unlike zymosan, which is a strong activator of mitogen-activated protein kinase (MAPK), ATP and PAF were weak MAPK activators and induced only a partial and transient increase in cPLA 2 phosphorylation (gel shift). However, ATP or PAF together with colony stimulating factor-1 (CSF-1) synergistically stimulated arachidonic acid release. CSF-1 is a strong MAPK activator that induces a rapid and complete cPLA 2 gel shift but not calcium mobilization or arachidonic acid release. Arachidonic acid release was more rapid in response to CSF-1 plus ATP or PAF than zymosan and correlated with the time course of the cPLA 2 gel shift. Although low concentrations of ionomycin induced a lower magnitude of calcium mobilization than ATP, the response was more sustained resulting in arachidonic acid release. A23187 and ionomycin induced weak MAPK activation, and a partial and transient cPLA 2 gel shift. The MAPK kinase inhibitor, PD 98059 suppressed A23187-induced MAPK activation and cPLA 2 gel shift but had little effect on arachidonic acid release. These results indicate that in macrophages a transient increase in [Ca 2؉ ] i and sustained phosphorylation of cPLA 2 can act together to promote arachidonic acid release but neither alone is sufficient. A sustained increase in calcium is sufficient for inducing arachidonic acid release. However, PMA and okadaic acid induce arachidonic acid release without increasing [Ca 2؉ ] i , although resting levels of calcium are required, suggesting alternative mechanisms of regulation.

Polychlorinated biphenyls induce arachidonic acid release in human platelets in a tamoxifen sensitive manner via activation of group IVA cytosolic phospholipase A2-α

“…calcium-dependent binding via the C2 domain [13][14][15] or calcium-independent binding through a phospholipidbinding region in the catalytic domain [21,19]. Furthermore, agents such as okadaic acid and phorbol myristate acetate (PMA) induces arachidonic acid release without increasing intracellular calcium concentration [22]. Also, it was reported earlier that thrombin-stimulated platelets produced more TXA 2 in the absence of extracellular calcium [64].…”

“…The catalytic domain can affect membrane association of the enzyme by modulating the rate of association and also the residence time at membranes [18]. Furthermore, there is now convincing evidence that calcium-independent pathways for cytosol-to-membrane translocation of cPLA 2 -α exists [19][20][21][22][23][24], thus, suggesting alternative mechanisms for membrane association. One such mechanism could involve calcium-independent association of cPLA 2 -α to anionic phospholipids [21,24,25].…”

Potentiation by vitamin D analogs of TNFα and ceramide-induced apoptosis in MCF-7 cells is associated with activation of cytosolic phospholipase A2

“…cPLA 2 is normally located in the cytoplasm and it translocates to the cell membrane in a calcium dependent or independent manner. 25,26 In order to define the cellular and molecular mechanisms involved in vitamin D-induced apoptosis in breast cancer cells, we have assessed the effects of two vitamin D analogs EB1089 and CB1093 on TNFa-induced apoptosis in MCF-7 cells. We now report that these vitamin D analogs potentiate responsiveness of breast cancer cells to TNFa and suggest that ceramide and/or cPLA 2 might be involved in a common pathway in TNFa and vitamin D mediated apoptosis.…”

Altered Endothelin-1 Signaling in Production of Thromboxane A2 in Kupffer Cells from Bile Duct Ligated Rats

“…Stimulation of Gi coupled receptors have been reported to lead to the phosphorylation of cPLA 2 and release of AA via the activation of p38 MAPK and ERK1/2, although the intermediates involved in the activation of p38 have not been fully elucidated (38)(39)(40)(41)(42)(43)(44)(45)(46)(47). KCs express ET (B) receptors coupled to Gq/G11 and Gi proteins (48)(49)(50)(51), and several reports have supported the initial findings by Qui et al of the requirements of both a transient increase in calcium and phosphorylation of ser505, via p38 MAPK or ERK1/2, for the full activation of cPLA 2 and AA release in macrophages (28,34,35).…”

“…This domain is a Ca 2+ and phospholipid binding domain that promotes interaction of cPLA 2 with its target membrane. Although the phosphorylation of Ser505 by p38 mitogen activated protein kinase (p38 MAPK) and extracellular signal-regulated kinase 1/2 (ERK1/2) (29-31), Ser515 by Ca 2+ /Calmodulin protein kinase II (CaMKII) (32), and Ser727 by MAPK interacting kinase I (Mnk1) (33) have been implicated in the activation of cPLA 2 ; several studies have established that the phosphorylation of Ser505 is critical for maximal activation of cPLA 2 in most cell types in response to agonist stimulation in vivo and in vitro (26,(29)(30)(31)(34)(35)(36)(37).…”