Characterization of the effects of Cl- channel modulators on TMEM16A and bestrophin-1 Ca2+ activated Cl- channels
Characterization of the effects of Cl- channel modulators on TMEM16A and bestrophin-1 Ca2+ activated Cl- channels
复制标题
Cl-通道调节剂对 TMEM16A 和 bestropin-1 Ca2 激活的 Cl-通道影响的表征
DOI：
10.1007/s00424-014-1572-5
复制
发表时间：
2015-07-01
影响因子：
4.5
通讯作者：
Zhang, Hailin
中科院分区：
文献类型：
--
作者：
Liu, Yani;Zhang, Huiran;Zhang, Hailin
钙激活的氯离子通道(CaCC)具有多种重要的生理功能。许多候选蛋白被提出形成CACC，但只有两个家族，bestrophins和TMEM16蛋白，在表达系统中概括了天然CACC的性质。由于缺乏特定的药理学，内源性CaCC的研究受到阻碍，因为大多数氯离子通道调节剂缺乏选择性，并且缺乏对这些调节剂对TMEM16A和Bestrophin的影响的系统比较。本研究采用膜片钳技术研究了七种氯离子通道阻滞剂尼氟米酸(NFA)、NPPB、氟苯那酸(FFA)、DIDS、单宁酸、CaCCinh-A01和T16A(Inh)-A01对中国仓鼠卵巢细胞稳定表达TMEM16A和Bestrophin-1(Best1)的影响。在所研究的7种缓蚀剂中，NFA对TMEM16A的选择性最高(IC50值为7.40+/-0.95mU M)，高于Best1(IC50值为102.19+/-15.05mU M)。相反，DIDS和TMEM16A对BEST1有反向选择性抑制作用，IC50值分别为3.93+/-0.73mU和548.86+/-25.57mU。CaCCinh-A01对TMEM16A和BEST1通道的阻断效果最好。T16A(Inh)-A01部分抑制TMEM16A电流，但对Best1电流无影响。单宁酸、NPPB和FFA有不同的中间效应。文中还描述了其中一些调节剂对通道活性的增强作用以及对TMEM16A失活动力学的影响。氯离子通道调节剂对TMEM16A和Best1的影响的表征将有助于未来对天然CaCC的研究。
The Ca2+ activated Cl- channels (CaCCs) play a multitude of important physiological functions. A number of candidate proteins have been proposed to form CaCC, but only two families, the bestrophins and the TMEM16 proteins, recapitulate the properties of native CaCC in expression systems. Studies of endogenous CaCCs are hindered by the lack of specific pharmacology as most Cl- channel modulators lack selectivity and a systematic comparison of the effects of these modulators on TMEM16A and bestrophin is missing. In the present study, we studied seven Cl- channel inhibitors: niflumic acid (NFA), NPPB, flufenamic acid (FFA), DIDS, tannic acid, CaCCinh-A01 and T16A(inh)-A01 for their effects on TMEM16A and bestrophin-1 (Best1) stably expressed in CHO (Chinese hamster ovary) cells using patch clamp technique. Among seven inhibitors studied, NFA showed highest selectivity for TMEM16A (IC50 of 7.40 +/- 0.95 mu M) over Best1 (IC50 of 102.19 +/- 15.05 mu M). In contrast, DIDS displayed a reverse selectivity inhibiting Best1 with IC50 of 3.93 +/- 0.73 mu M and TMEM16A with IC50 of 548.86 +/- 25.57 mu M. CaCCinh-A01 was the most efficacious blocker for both TMEM16A and Best1 channels. T16A(inh)-A01 partially inhibited TMEM16A currents but had no effect on Best1 currents. Tannic acid, NPPB and FFA had variable intermediate effects. Potentiation of channel activity by some of these modulators and the effects on TMEM16A deactivation kinetics were also described. Characterization of Cl- channel modulators for their effects on TMEM16A and Best1 will facilitate future studies of native CaCCs.