Mutation in a protein kinase C phosphorylation site of the 5-HT1A receptor preferentially attenuates Ca2+ responses to partial as opposed to higher-efficacy 5-HT1A agonists
Mutation in a protein kinase C phosphorylation site of the 5-HT1A receptor preferentially attenuates Ca2+ responses to partial as opposed to higher-efficacy 5-HT1A agonists
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DOI：
10.1016/s0028-3908(03)00097-2
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发表时间：
2003-06-01
影响因子：
4.7
通讯作者：
Pauwels, PJ
中科院分区：
文献类型：
--
作者：
Wurch, T;Colpaert, FC;Pauwels, PJ
在5-HT 1A受体的第二胞内环的推定蛋白激酶C磷酸化位点中的Thr（149）Ala突变已显示影响Ca 2+通道的关闭和Ca 2+动员，而不干扰抑制性cAMP途径（Mol Pharmacol 52（1997）164）。这里，在野生型（wt）和突变型Thr（149）Ala 5-HT 1A受体之间比较了一系列5-HT 1A激动剂的Ca 2+响应，所述受体作为含有G（α 15）蛋白的融合蛋白的一部分。与野生型5-HT 1A受体蛋白相比，突变和融合过程都没有改变融合蛋白的基于[H-3]WAY 100635的配体结合谱。然而，在野生型5-HT 1A受体，5-HT诱导的Ca 2+反应在CHO-K1细胞通过内源性G（i/o）蛋白，Ca 2+反应5-HT在突变体Thr（149）Ala 5-HT 1A受体是完全依赖于共表达或融合到重组G（α 15）蛋白。丁螺环酮、flesinoxan和8-OH-DPAT在wt 5-HT 1A：G（alpha 15）融合蛋白处产生分级的部分响应（26至62%）; F 13640、5-CT和F 14679表现为具有与5-HT类似的最大Ca 2+响应的更高功效的激动剂。对于flesinoxan和8-OH-DPAT，突变体Thr（149）Ala 5-HT 1A：G（alpha 15）融合蛋白的最大Ca 2+反应显著减弱（分别为-45和-36%）;对其他5-HT激动剂的反应没有显著影响。在用佛波醇12-肉豆蔻酸酯13-乙酸酯处理Thr（149）Ala 5-HT 1A：G（α 15）融合蛋白时观察到类似的效果。总之，由部分而非完全5-HT 1A受体激动剂诱导的Ca 2+反应的幅度受5-HT 1A：G（α 15）融合蛋白的Thr（149）Ala突变的影响。(C)2003爱思唯尔科技有限公司版权所有。
The Thr(149)Ala mutation in a putative protein kinase C phosphorylation site of the 5-HT1A receptor's second intracellular loop has been shown to affect the closing of Ca2+ channels and Ca2+ mobilisation without interfering with the inhibitory cAMP pathway (Mol Pharmacol 52 (1997) 164). Here, the Ca2+ responses for a series of 5-HTIA agonists were compared between the wild-type (wt) and mutant Thr(149)Ala 5-HT1A receptor as part of a fusion protein containing a G(alpha15) protein. Neither the mutation nor the fusion process modified the [H-3]WAY 100635-based ligand binding profile of the fusion proteins as compared to the wt 5-HT1A receptor protein. Whereas at the wt 5-HT1A receptor, 5-HT induced a Ca2+ response in CHO-K1 cells via endogenous G(i/o) proteins, the Ca2+ response to 5-HT at the mutant Thr(149)Ala 5-HT1A receptor was fully dependent on either the co-expression or the fusion to a recombinant G(alpha15) protein. Buspirone, flesinoxan and 8-OH-DPAT produced a graded partial response (26 to 62%) at the wt 5-HT1A:G(alpha15) fusion protein; F 13640, 5-CT and F 14679 behaved as higher-efficacy agonists with maximal Ca2+ responses similar to 5-HT. The maximal Ca2+ responses at the mutant Thr(149)Ala 5-HT1A:G(alpha15) fusion protein were significantly attenuated for flesinoxan and 8-OH-DPAT (-45 and -36%, respectively); the response to the other 5-HT agonists was not significantly affected. A similar effect was observed upon treatment with phorbol 12-myristate 13-acetate at the Thr(149)Ala 5-HT1A:G(alpha15) fusion protein. In conclusion, the amplitude of the Ca2+ responses induced by partial, but not that to fuller 5-HT1A receptor agonists, is affected by the Thr(149)Ala mutation of the 5-HT1A:G(alpha15) fusion protein. (C) 2003 Elsevier Science Ltd. All rights reserved.