调控T型钙通道Ca_v3.1电压依赖性失活的分子结构域(英文)

Molecular Regions Determining The Voltage-dependence of Inactivation for T-type Calcium Channel Ca_v3.1

T型钙通道（Cav3）广泛分布于各类细胞,其显著的电生理学特点是低电位激活和快速的电压依赖性失活.失活在通道的生理功能调节中起十分重要的作用,但具体参与通道失活的分子基础目前并不完全清楚.为明确Cav3.1通道中调控电压依赖性失活的结构域,用Cav1.2通道（无电压依赖性失活）结构域Ⅰ和Ⅱ中的S1-S4、S5-S6区及Ⅰ和Ⅱ间的联系区替换Cav3.1中的相应区域,构建嵌合通道,并在卵母细胞中表达,用电压钳技术分析通道的电生理学特性.结果表明,替换Ⅰ中的S1-S4或S5-S6区可使Cav3.1的失活特性显著改变,但这种改变主要是由激活-失活偶联所致.Ⅱ的替换使通道的失活曲线参数发生显著改变,表明结构域Ⅱ,包括S1-S4和S5-S6均参与Cav3.1失活过程的调控.Ⅰ、Ⅱ间的联系区及Ⅰ中的S5-S6主要调控Cav3.1的失活速率,Ⅰ和Ⅱ中的S1-S4对通道失活速率无影响.综上所述,结构域Ⅱ是调控Cav3.1电压依赖性失活的关键因素,结构域Ⅰ不参与该通道失活过程的调控.Ⅰ、Ⅱ间的联系区及Ⅰ中的S5-S6主要调控Cav3.1通道的失活速率,Ⅰ、Ⅱ中的S1-S4对通道失活速率无影响.

The notable features for inactivation of Cav3 channels are fast inactivating rate and strong voltage-dependence. We have investigated the molecular basis for determining the voltage-dependence of inactivation for Cav3.1, focusing on domain Ⅰ and Ⅱ. We made chimeras between Cav3.1 and Cav.2. Chimeras were expressed in oocytes and currents were recorded by voltage clamp. For domain Ⅰ, replacement of S1 -S4 or S5-S6 shifted the steady state inactivation curve significantly. These changes were mainly or partially caused by activation-inactivation coupling, rather than molecular modification. Replacement of domain Ⅱ shifted the inactivation curve significantly and these changes refer to molecular modification, indicating that domain Ⅱ contributed to the voltage-dependence of inactivation for Ca,3.1. Furthermore, both voltage sensor region S 1 - S4 and pore region S5 - S6 in domain Ⅱ were also involved, but Ⅰ - Ⅱ linker has no contribution. In addition, we found that the Ⅰ - Ⅱ linker and S5 -S6 in domain Ⅰ contributed strongly to inactivation rate for Cav3.1, while S 1 - S4 in domain Ⅰ and Ⅱ was not involved. Taken collectively, our results suggest that domain Ⅱ plays a key role in determining the voltage-dependence of inactivation for Ca,3.1, which was different from the molecular determinants for inactivating rate and for voltage-dependence of activation.