Denys Anne, Aires Virginie, Hichami Aziz, Khan Naim Akhtar
Département de Physiologie, UPRES Lipides et Nutrition, Université de Bourgogne, Faculté des Sciences de la Vie, 6 Boulevard Gabriel, 21000 Dijon, France.
FEBS Lett. 2004 Apr 23;564(1-2):177-82. doi: 10.1016/S0014-5793(04)00361-8.
This study was conducted on human Jurkat T-cells to investigate the role of depletion of intracellular Ca(2+) stores in the phosphorylation of two mitogen-activated protein kinases (MAPKs), i.e. extracellular signal-regulated kinase (ERK) 1 and ERK2, and their modulation by a polyunsaturated fatty acid, docosahexaenoic acid (DHA). We observed that thapsigargin (TG) stimulated MAPK activation by store-operated calcium (SOC) influx via opening of calcium release-activated calcium (CRAC) channels as tyrphostin-A9, a CRAC channel blocker, and two SOC influx inhibitors, econazole and SKF-96365, diminished the action of the former. TG-stimulated ERK1/ERK2 phosphorylation was also diminished in buffer containing EGTA, a calcium chelator, further suggesting the implication of calcium influx in MAPK activation in these cells. Moreover, TG stimulated the production of diacylglycerol (DAG) by activating phospholipase D (PLD) as propranolol (PROP) (a PLD inhibitor), but not U73122 (a phospholipase C inhibitor), inhibited TG-evoked DAG production in these cells. DAG production and protein kinase C (PKC) activation were involved upstream of MAPK activation as PROP and GF109203X, a PKC inhibitor, abolished the action of TG on ERK1/ERK2 phosphorylation. Furthermore, DHA seems to act by inhibiting PKC activation as this fatty acid diminished TG- and phorbol 12-myristate 13-acetate-induced ERK1/ERK2 phosphorylation in these cells. Together these results suggest that Ca(2+) influx via CRAC channels is implicated in PLD/PKC/MAPK activation which may be a target of physiological agents such as DHA.
本研究以人Jurkat T细胞为对象，旨在探究细胞内Ca(2+)储存耗竭在两种丝裂原活化蛋白激酶（MAPK），即细胞外信号调节激酶（ERK）1和ERK2磷酸化过程中的作用，以及多不饱和脂肪酸二十二碳六烯酸（DHA）对其的调节作用。我们观察到，毒胡萝卜素（TG）通过打开钙释放激活钙（CRAC）通道，经储存操纵性钙（SOC）内流刺激MAPK激活，因为CRAC通道阻滞剂 tyrphostin - A9以及两种SOC内流抑制剂酮康唑和SKF - 96365可减弱前者的作用。在含有钙螯合剂乙二醇双四乙酸（EGTA）的缓冲液中，TG刺激的ERK1/ERK2磷酸化也减弱，这进一步表明钙内流与这些细胞中MAPK激活有关。此外，TG通过激活磷脂酶D（PLD）刺激二酰基甘油（DAG）的产生，因为普萘洛尔（PROP）（一种PLD抑制剂）而非U73122（一种磷脂酶C抑制剂）可抑制这些细胞中TG诱发的DAG产生。DAG产生和蛋白激酶C（PKC）激活参与MAPK激活的上游过程，因为PROP和PKC抑制剂GF109203X可消除TG对ERK1/ERK2磷酸化的作用。此外，DHA似乎通过抑制PKC激活发挥作用，因为这种脂肪酸可减弱这些细胞中TG和佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯诱导的ERK1/ERK2磷酸化。这些结果共同表明，通过CRAC通道的Ca(2+)内流与PLD/PKC/MAPK激活有关，而这可能是DHA等生理活性剂的作用靶点。
