P2X7 receptor-Pannexin1 complex:: pharmacology and signaling
P2X7 receptor-Pannexin1 complex:: pharmacology and signaling
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DOI：
10.1152/ajpcell.00228.2008
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发表时间：
2008-09-01
影响因子：
5.5
通讯作者：
Scemes, E.
中科院分区：
文献类型：
--
作者：
Iglesias, R.;Locovei, S.;Scemes, E.
Pannexin 1（Panx 1）是无脊椎动物连接蛋白间隙连接的直系同源物，最近被认为是P2 X（7）受体（P2 X（7）R）激活诱导的孔。我们探索了已知阻断P2 X（7）R激活的Panx 1通道上间隙连接的化合物的药理作用以及这两种蛋白质之间相互作用的机制。全细胞记录显示不同的P2 X（7）R和Panx 1电流响应激动剂。P2 X（7）R刺激或膜去极化后Panx 1电流的激活被Panx 1小干扰RNA（siRNA）阻断，并被甲氟喹>甘珀酸>氟灭酸阻断。用KN-62（一种P2 X（7）R拮抗剂）孵育细胞，可阻止2 '（3'）O-（4-苯甲酰基苯甲酰基）腺苷5 '-三磷酸（BzATP）激活电流。Panx 1 siRNA和甘珀酸和甲氟喹也阻止了BzATP诱导的染料膜透化。膜渗透性（TAT-P2 X（7））肽提供的证据表明，P2 X（7）R的COOH末端的Src同源性3死亡结构域参与导致Panx 1活化的信号转导事件的初始步骤，并且Src酪氨酸激酶可能参与该过程。竞争分析表明，20 μ M TAT-P2 X（7）肽导致Src与P2 X（7）R复合物结合减少50%。KN-62、TAT-P（2）X7肽和Src酪氨酸抑制剂PP 2降低了BzATP刺激后Src酪氨酸磷酸化，这些化合物阻止了大电导Panx 1电流和膜透化。这些结果以及P2 X（7）R刺激后Panx 1酪氨酸磷酸化的缺失表明，在通过P2 X（7）R介导Panx 1活化的酪氨酸激酶信号转导途径中，涉及了另外的分子。
Pannexin 1 (Panx1), an ortholog to invertebrate innexin gap junctions, has recently been proposed to be the pore induced by P2X(7) receptor (P2X(7)R) activation. We explored the pharmacological action of compounds known to block gap junctions on Panx1 channels activated by the P2X(7)R and the mechanisms involved in the interaction between these two proteins. Whole cell recordings revealed distinct P2X(7)R and Panx1 currents in response to agonists. Activation of Panx1 currents following P2X(7)R stimulation or by membrane depolarization was blocked by Panx1 small-interfering RNA (siRNA) and with mefloquine > carbenoxolone > flufenamic acid. Incubation of cells with KN-62, a P2X(7)R antagonist, prevented current activation by 2'(3')O-( 4-benzoylbenzoyl) adenosine 5'-triphosphate (BzATP). Membrane permeabilization to dye induced by BzATP was also prevented by Panx1 siRNA and by carbenoxolone and mefloquine. Membrane permeant (TAT-P2X(7)) peptides, provided evidence that the Src homology 3 death domain of the COOH-terminus of the P2X(7)R is involved in the initial steps of the signal transduction events leading to Panx1 activation and that a Src tyrosine kinase is likely involved in this process. Competition assays indicated that 20 mu M TAT-P2X(7) peptide caused 50% reduction in Src binding to the P2X(7)R complex. Src tyrosine phosphorylation following BzATP stimulation was reduced by KN-62, TAT-P(2)X7 peptide, and by the Src tyrosine inhibitor PP2 and these compounds prevented both large-conductance Panx1 currents and membrane permeabilization. These results together with the lack Panx1 tyrosine phosphorylation in response to P2X(7)R stimulation indicate the involvement of an additional molecule in the tyrosine kinase signal transduction pathway mediating Panx1 activation through the P2X(7)R.