Diacylglycerol lipase activation and 5-lipoxygenase activation and translocation following TCR/CD3 triggering in T cells.
Arachidonic acid (AA) release was observed following T cell receptor (TCR)/CD3 complex cross-linking in different tumor T cell lines as well as on purified peripheral T cells in vivo. Direct measurement of enzymatic activity in vitro of TCR/CD3-stimulated Jurkat cell extracts on labeled vesicle substrates showed that TCR/CD3 cross-linking resulted in AA release from sn-1,2-diacylglycerol (DAG) vesicles, as detected by TLC analysis, suggesting that DAG lipase was activated following TCR/CD3 stimulation and DAG generation. On the contrary, no phospholipase A2 activation was observed in response to TCR/CD3 stimulation, since no lyso-phospholipids were generated in vitro from either phosphatidylcholine or phosphatidylinositol-3,4-bisphosphate, or from phosphatidic acid vesicles. Moreover, the 1-DAG lipase inhibitor RHC80267 completely blocked TCR/CD3-dependent AA release in vitro and in vivo, without effect upon TCR/CD3-dependent inositol-1,4,5-trisphosphate (IP3) generation. Importantly, evidence for further metabolism of released AA was obtained, since synthesis and release of cysteinyl leukotrienes (CLT), but not of leukotriene B4 or cyclooxygenase products, could be detected by radioimmunoassay in different T cell lines and peripheral blood T cells following TCR/CD3 cross-linking. Moreover, HPLC analysis revealed an accumulation of leukotriene E4 in TCR/CD3 stimulated Jurkat cells. This was associated with translocation of 5-lipoxygenase from the cytosol to the cell membranes. Finally, TCR/CD3-mediated CLT production was blocked by MK886, a specific inhibitor of 5-LO translocation and activation. Our data help define a further level in the fate of second messengers generated after TCR/CD3 triggering and suggest that additional mediators can play a role in the context of T cell activation.
Cifone
M G
MG
Department of Experimental Medicine, University of L'Aquila, Italy.
Cironi
L
L
Santoni
A
A
Testi
R
R
D001094:Arachidonate 5-Lipoxygenase / Q000096:biosynthesis*; D017252:CD3 Complex / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D008071:Lipoprotein Lipase / Q000096:biosynthesis*; D008213:Lymphocyte Activation; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D015290:Second Messenger Systems; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000378:metabolism*
D017252:CD3 Complex; D011948:Receptors, Antigen, T-Cell; D001094:Arachidonate 5-Lipoxygenase; D008071:Lipoprotein Lipase