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The roles of calpastatin and calmodulin in the regulation of calcium channel

钙蛋白酶抑制剂和钙调蛋白在钙通道调节中的作用

基本信息

批准号：
15570137
负责人：
HAO Li-ying
金额：
$ 1.34万
依托单位：
Kagoshima University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15570137/
关键词：
calmodulin calpastatin CaMKII Ca^<2+> channel run-down CaMK Ca^<2+> channel

项目摘要

Voltage-gated L-type Ca^<2+> channels control diverse physiological functions including gene transcription, rhythmic firing, synaptic transmission, hormone secretion and excitation-contraction coupling. Activity of L-type Ca^<2+> channel decreases when the cytoplasmic side of the channel is perfused with an artificial physiological solution (run-down phenomenon). The mechanism of run-down is still unknown. We have previously reported that both calpastatin(CS) and calmodulin(CaM) can recover Ca^<2+> channel activity after run-down. In this study, we have investigated the relations and the mechanisms of calmodulin and calpastatin in the activation of Ca^<2+> channel and the reversion of run-down. The main results are as follows. (1)The run-down reversing effect of CaM depends on channel phosphorylation state. CaM kinase II keeps the channel in a state that can be reactivated by CaM. (2)Domain L of CS reactivates the L-type Ca^<2+> channel. By screening the short peptides of domain L, we found that the effective region of CS is located in the amino acid residues 45-64 of domain L. (3)To investigate the relations of CS and CaM, binding between these two molecules were examined with GST-fusion protein pull-down assay. The result shows that CS and CaM bind each other in the presence of Ca^<2+>. (4)Binding sites of CS to Ca^<2+> channel were also investigated. CS might bind to the region that has already confirmed as CaM binding region.

电压门控的l型Ca^<2+>通道控制着多种生理功能，包括基因转录、节律放电、突触传递、激素分泌和兴奋-收缩耦合。当通道细胞质侧灌注人工生理溶液时，l型Ca^<2+>通道活性降低（损耗现象）。衰竭的机制尚不清楚。我们之前报道过钙pastatin（CS）和calmodulin（CaM）都可以在运行后恢复Ca^<2+>通道活性。在本研究中，我们研究了钙调素和钙pastatin在激活Ca^<2+>通道和逆转rundown中的关系和机制。主要结果如下：(1) CaM的运行逆转作用取决于通道磷酸化状态。CaM激酶II使通道处于可被CaM重新激活的状态。(2) CS的L域重新激活了L型Ca^<2+>通道。通过筛选L结构域短肽，我们发现CS的有效区域位于L结构域45-64个氨基酸残基上。(3)为了研究CS和CaM的关系，我们采用gst融合蛋白下拉实验检测了这两个分子之间的结合。结果表明，CS和CaM在Ca^<2+>存在下相互结合。(4) CS与Ca^<2+>通道的结合位点也进行了研究。CS可能会绑定到已经确认为CaM绑定区的区域。