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首页> 外文期刊>Cellular Physiology and Biochemistry >Homologous desensitisation of the mouse leukotriene B-4 receptor involves protein kinase C-mediated phosphorylation of serine 127
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Homologous desensitisation of the mouse leukotriene B-4 receptor involves protein kinase C-mediated phosphorylation of serine 127

机译:小鼠白三烯B-4受体的同源脱敏涉及蛋白激酶C介导的丝氨酸127磷酸化

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Murine leukotriene B-4 (LTB4) receptor (mBLT1) cDNA was identified by searching the EST database using human LTB4 receptor as the query sequence. Expression of functional mBLT1 after injection of in vitro transcribed cRNA into Xenopus laevis oocytes was demonstrated as LTB4-evoked, Ca2+-activated Cl(-)currents recorded by two-electrode voltage clamp. From mBLT1-expressing oocytes, a dose-dependent relationship between the Ca2+-activated Cl- current and LTB4 concentration was demonstrated with an apparent EC50 of 6.7 nM. Following LTB4 stimulation of mBLT1, we observed two transient, spatially distinct Ca2+-activated, inwardly directed Cl- currents in the oocytes: a fast peak current requiring relatively high LTB4 concentrations, and a slowly progressing Cl(-)current. Nucleotides, PGE(2), 12R-hydroxy-5, 8, 14-cis10-trans-eicosatetraenoic acid, and LTD4 did not activate mBLT1. U75302, specifically targeting BLT1, significantly reduced LTB4- evoked Cl- currents. Repetitive LTB4 administration desensitized the LTB 4evoked currents. Activation of protein kinase C (PKC) by PMA addition completely eliminated the LTB(4)evoked currents, whereas down-regulation of PKC by prolonged PMA exposure (20 h) impaired mBLT1 desensitisation. In addition, Ser-127-Ala substitution of the PKC consensus phosphorylation site on the second intracellular loop prevented the mBLT1 desensitisation. These data indicate that PKCmediated phosphorylation at Ser-127 leads to mBLT1 desensitisation. Copyright (c) 2007 S. Karger AG, Basel.
机译:鼠白三烯B-4(LTB4)受体(mBLT1)cDNA是通过使用人LTB4受体作为查询序列搜索EST数据库来鉴定的。体外转录的cRNA注射到非洲爪蟾卵母细胞中后,功能性mBLT1的表达被LTB4诱发,Ca2 +激活的Cl(-)电流通过两电极电压钳记录。从表达mBLT1的卵母细胞中,Ca2 +激活的Cl-电流与LTB4浓度之间存在剂量依赖性关系,表观EC50为6.7 nM。继LTB4刺激mBLT1之后,我们观察到卵母细胞中两个瞬时的,空间上不同的Ca2 +激活的,向内定向的Cl-电流:需要相对较高LTB4浓度的快速峰电流和缓慢进展的Cl(-)电流。核苷酸,PGE(2),12R-羟基-5,8,14-cis10-反式二十碳四烯酸和LTD4不会激活mBLT1。 U75302(特别针对BLT1)显着降低了LTB4诱发的Cl电流。重复进行LTB4管理会使LTB 4诱发的电流失去敏感性。 PMA的添加激活蛋白激酶C(PKC)完全消除了LTB(4)引起的电流,而长时间PMA暴露(20 h)导致PKC的下调削弱了mBLT1脱敏。此外,第二个细胞内环上的PKC共有磷酸化位点的Ser-127-Ala取代阻止了mBLT1脱敏。这些数据表明,在Ser-127处PKC介导的磷酸化导致mBLT1脱敏。版权所有(c)2007 S.Karger AG,巴塞尔。

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