Functional expression of TrpC1: a human homologue of the Drosophila Trp channel.
Functional expression of TrpC1: a human homologue of the Drosophila Trp channel.
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DOI：
10.1042/bj3310331
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发表时间：
1998-04
期刊：
影响因子：
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通讯作者：
W. Sinkins;M. Estacion;W. Schilling
中科院分区：
文献类型：
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作者：
W. Sinkins;M. Estacion;W. Schilling
TrpC1在哺乳动物细胞中表达时，似乎是一个存储操作通道（SOC）。在本研究中，TrpC1在Sf9昆虫细胞中使用杆状病毒表达系统表达。TrpC1的表达引起基础胞浆游离Ca 2+浓度（[Ca 2 +]i）的增加作为感染后时间的函数。基础的Ba2+内流，质膜Ca2+渗透性的指数，也增加，并被封锁的La 3+。虽然毒胡萝卜素诱导的[Ca2+]i的变化在TrpC1表达细胞中比对照组更大，但Ba2+内流不受毒胡萝卜素的影响。在TrpC1表达细胞中记录的全细胞膜电流随着感染后时间的增加而增加，并且（1）在对称的葡萄糖酸钠溶液中向内整流，（2）对Na+、Ca2+和Ba2+无选择性，（3）被La3+阻断。此外，TrpC 1电流不受（1）毒胡萝卜素、（2）用Ins（1，4，5）P3透析细胞或（3）用含有高浓度Ca 2+螯合剂EGTA的溶液透析细胞的影响。这些结果表明，TrpC1形成非选择性阳离子通道，当在Sf9细胞中表达时，其是组成性活性的，但对内部Ca2+储存的耗尽不敏感。因此，TrpCl可能是SOC的亚基，其单独可以在Sf9细胞中形成功能通道，但是其需要存在于哺乳动物细胞中的另外的亚基或细胞质因子来表达SOC活性。
TrpC1 appears to be a store-operated channel (SOC) when expressed in mammalian cells. In the present study, TrpC1 was expressed in Sf9 insect cells using the baculovirus expression system. Expression of TrpC1 caused an increase in basal cytosolic free Ca2+ concentration ([Ca2+]i) as a function of post-infection time. Basal Ba2+ influx, an index of plasmalemmal Ca2+ permeability, was also increased and was blocked by La3+. Although the thapsigargin-induced change in [Ca2+]i was greater in TrpC1-expressing cells than controls, Ba2+ influx was unaffected by thapsigargin. Whole-cell membrane currents recorded in TrpC1-expressing cells increased as a function of post-infection time and were (1) inwardly rectifying in symmetrical sodium gluconate solutions, (2) non-selective with respect to Na+, Ca2+ and Ba2+, and (3) blocked by La3+. Furthermore TrpC1 currents were unaffected by (1) thapsigargin, (2) dialysis of the cell with Ins(1,4,5)P3 or (3) dialysis of the cell with solutions containing high concentrations of the Ca2+ chelator, EGTA. These results suggest that TrpC1 forms non-selective cation channels that are constitutively active when expressed in Sf9 cells, but insensitive to depletion of the internal Ca2+ stores. Thus TrpC1 may be a subunit of a SOC which alone can form functional channels in Sf9 cells, but which requires additional subunits or cytoplasmic factors present in mammalian cells for expression of SOC activity.