Crucial role of N terminus in function of cardiac L-type Ca2+ channel and its modulation by protein kinase C
Crucial role of N terminus in function of cardiac L-type Ca2+ channel and its modulation by protein kinase C
复制标题
DOI：
10.1074/jbc.273.28.17901
复制
发表时间：
1998-07-10
影响因子：
4.8
通讯作者：
Dascal, N
中科院分区：
文献类型：
--
作者：
Shistik, E;Ivanina, T;Dascal, N
研究了心脏l型电压依赖性Ca2+通道主要亚基（α (1C)）胞浆N端在非洲爪蟾卵母细胞表达系统中的作用。如先前报道（Wei， X., Neely, a., Olcese， R., Lang， W., Stefani， E., and Birnbaumer， L.(1996)），兔心脏α (1C)中最初的46或139个氨基酸（a.a.）的缺失会导致Ba2+ (I-Ba)携带的全细胞Ca2+通道电流增加5-10倍。通道4,205-215)，免疫化学测量的α (1C)蛋白质膜含量未被46-a.a a改变。删除。在二氢吡啶激动剂存在的情况下膜片钳记录显示，这种缺失导致单通道打开概率增加5-10倍，而不改变通道密度。因此，N端初始段影响通道门控而不是表达。辅助β (2A)亚基的共表达引起的I-Ba的增加在全长α (1C)通道中比在46-或139-a.a通道中明显增强。截断突变体，表明β (2A)和N端之间存在相互作用。然而，在体外，只有α的I-II结构域连接体（1C）结合β (2A)，而不是与N或C末端结合的α - ii结构域连接体（2A）。删除。因此，α (1C)的N端在通道门控和PKC调制中起着至关重要的作用。我们认为PKC和β亚基通过减轻N端施加的抑制控制来增强通道的活性。由于PKC上调l型Ca2+通道在许多物种中都有报道，我们预测含有初始n端46 a.a.的α (1C)亚基同工型与兔心脏α (1C)相似，广泛存在于心脏和平滑肌细胞中。
The role of the cytosolic N terminus of the main subunit (alpha(1C)) of cardiac L-type voltage-dependent Ca2+ channel was studied in Xenopus oocyte expression system. Deletion of the initial 46 or 139 amino acids (a.a.) of rabbit heart alpha(1C) caused a 5-10-fold increase in the whole cell Ca2+ channel current carried by Ba2+ (I-Ba), as reported previously (Wei, X., Neely, A., Olcese, R., Lang, W., Stefani, E., and Birnbaumer, L. (1996) Recept. Channels 4, 205-215), The plasma membrane content of alpha(1C) protein, measured immunochemically, was not altered by the 46-a.a. deletion. Patch clamp recordings in the presence of a dihydropyridine agonist showed that this deletion causes a 5-10-fold increase in single channel open probability without changing channel density. Thus, the initial segment of the N terminus affects channel gating rather than expression. The increase in I-Ba caused by coexpression of the auxiliary beta(2A) subunit was substantially stronger in channels with full-length alpha(1C) than in 46- or 139-a.a. truncated mutants, suggesting an interaction between beta(2A) and N terminus. However, only the I-II domain linker of alpha(1C), but not to N or C termini, bound beta(2A) in vitro, The well documented increase of I-Ba caused by activation of protein kinase C (PKC) was fully eliminated by the 46-a.a. deletion. Thus, the N terminus of alpha(1C) plays a crucial role in channel gating and PKC modulation, We propose that PKC and beta subunit enhance the activity of the channel in part by relieving an inhibitory control exerted by the N terminus. Since PKC up-regulation of L-type Ca2+ channels has been reported in many species, we predict that isoforms of alpha(1C) subunits containing the initial N-terminal 46 a.a. similar to those of the rabbit heart alpha(1C) are widespread in cardiac and smooth muscle cells.