Extracellular ATP‐mediated phospholipase a2 activation in rat thyroid FRTL‐5 cells: Regulation by a Gi/Go protein, Ca2+, and mitogen‐activated protein kinase
Extracellular ATP‐mediated phospholipase a2 activation in rat thyroid FRTL‐5 cells: Regulation by a Gi/Go protein, Ca2+, and mitogen‐activated protein kinase
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DOI：
10.1002/(sici)1097-4652(200005)183:2
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发表时间：
2000-05
影响因子：
5.6
通讯作者：
E. Ekokoski;B. Dugué;M. Vainio;P. Vainio;K. Törnquist
中科院分区：
文献类型：
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作者：
E. Ekokoski;B. Dugué;M. Vainio;P. Vainio;K. Törnquist
我们研究了大鼠甲状腺FRTL-5细胞中P2受体激动剂ATP激活磷脂酶A2（PLA 2）的机制。通过测量[3 H]-花生四烯酸（AA）从预标记细胞中的释放来测定PLA 2活性。ATP诱发剂量和时间依赖性AA释放。这种释放被百日咳毒素（PTX）处理完全抑制，表明Gi/Go蛋白的参与。用EGTA螯合细胞外Ca ~（2+）或用Ni ~（2+）抑制Ca ~（2+）内流也可减少AA的释放。虽然单独使用12-佛波醇13-肉豆蔻酸酯乙酸酯（PMA）激活蛋白激酶C（PKC）不会诱导任何AA释放，但当PKC被GF 109203 X抑制或与PMA长时间孵育以下调PKC时，ATP诱发的AA释放显著减少。ATP诱发的AA释放和丝裂原活化蛋白激酶（MAP激酶）磷酸化均被MAP激酶激酶（MEK）抑制剂PD 98059降低。此外，ATP诱发的MAP激酶磷酸化也被GF 109203 X和PKC下调抑制，表明PKC介导的MAP激酶活化。通过PP 1抑制Src样激酶减弱MAP激酶磷酸化和AA释放。这些结果表明，这些激酶参与MAP激酶和PLA 2激活的调节。TSH或dBucAMP引起的细胞内cAMP升高不诱导MAP激酶磷酸化。此外，TSH或dBucAMP均未减弱ATP诱发的AA释放和MAP激酶磷酸化。总之，我们的结果表明，ATP通过Gi/Go蛋白依赖性机制调节PLA 2的活化。此外，Ca 2+、PKC、MAP激酶和Src样激酶也参与了这一调节过程。J.细胞。183：155-162，2000。© 2000 Wiley利斯公司
We investigated the mechanism of phospholipase A2 (PLA2) activation in response to the P2 receptor agonist ATP in rat thyroid FRTL‐5 cells. The PLA2 activity was determined by measuring the release of [3H]‐arachidonic acid (AA) from prelabeled cells. ATP evoked a dose‐ and time‐dependent AA release. This release was totally inhibited by pertussis toxin (PTX) treatment, indicating the involvement of a Gi/Go protein. The AA release was also diminished by chelating extracellular Ca2+ with EGTA or by inhibiting influx of Ca2+ using Ni2+. Although the activation of protein kinase C (PKC) by 12‐phorbol 13‐myristate acetate (PMA) alone did not induce any AA release, the ATP‐evoked AA release was significantly reduced when PKC was inhibited by GF109203X or by a long incubation with PMA to downregulate PKC. Both the ATP‐evoked AA release and the mitogen‐activated protein kinase (MAP kinase) phosphorylation were decreased by the MAP kinase kinase (MEK) inhibitor PD98059. Furthermore, the ATP‐evoked MAP kinase phosphorylation was also inhibited by GF109203X and by downregulation of PKC, suggesting a PKC‐mediated activation of MAP kinase. Inhibiting Src‐like kinases by PP1 attenuated both the MAP kinase phosphorylation and the AA release. These results suggest that these kinases are involved in the regulation of MAP kinase and PLA2 activation. Elevation of intracellular cAMP by TSH or by dBucAMP did not induce a phosphorylation of MAP kinase. Furthermore, neither the ATP‐evoked AA release nor the MAP kinase phosphorylation were attenuated by TSH or dBucAMP. Taken together, our results suggest that ATP regulates the activation of PLA2 by a Gi/Go protein‐dependent mechanism. Moreover, Ca2+, PKC, MAP kinase, and Src‐like kinases are also involved in this regulatory process. J. Cell. Physiol. 183:155–162, 2000. © 2000 Wiley‐Liss, Inc.