Disruption of excitation–contraction coupling and titin by endogenous Ca2+‐activated proteases in toad muscle fibres
Disruption of excitation–contraction coupling and titin by endogenous Ca2+‐activated proteases in toad muscle fibres
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蟾蜍肌纤维中内源性 Ca2+ 激活蛋白酶对兴奋-收缩耦合和肌联蛋白的破坏
DOI：
10.1113/jphysiol.2004.082180
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
G. Lamb
中科院分区：
文献类型：
--
作者：
E. Verburg;R. Murphy;D. Stephenson;G. Lamb
本研究研究了细胞内游离[Ca 2 +]的生理水平升高对去极化诱导力反应的影响，以及对蟾蜍髂腓肌机械剥皮纤维中收缩装置产生的被动和主动力的影响。剥皮后保留了兴奋-收缩（EC）偶联，并且可以通过横管（T-）系统的去极化引起力反应。将细胞质[Ca 2 +]升高至≤ 1 μm或以上3 min，通过中断T系统中电压传感器与肌浆网中Ca 2+释放通道之间的耦合，导致去极化诱导力反应不可逆降低。这种解偶联显示出陡峭的[Ca 2 +]依赖性，在1.9 μm Ca 2+处解偶联50%。钙蛋白酶抑制剂亮抑酶肽（leupeptin）（1 mm）在很大程度上阻止了2 μm Ca ~（2+）引起的解偶联。将细胞质[Ca 2 +]升高到1 μm以上也会导致拉伸皮肤纤维中被动力产生的不可逆下降，其下降方式由[Ca 2 +]分级，尽管相对速度比耦合损失慢得多。将[Ca ~（2+）]降低至10 nm可迅速阻止被动力的逐渐丧失，1 mm亮抑酶肽几乎完全抑制被动力的逐渐丧失，而10 μm钙蛋白酶抑制剂则不能抑制被动力的逐渐丧失。经Ca 2+暴露预活化的肌肉匀浆也明显含有可扩散因子，其以Ca 2+依赖性方式对被动力产生造成损害。Western blotting显示：（a）钙蛋白酶-3存在于皮肤纤维中，并被Ca 2+暴露激活，（B）拉伸皮肤纤维中的Ca 2+暴露导致肌联蛋白的蛋白水解。我们得出结论，在[Ca 2 +]水平升高时发生的EC偶联的破坏可能至少部分由Ca 2+激活的蛋白酶引起，最有可能是由钙蛋白酶-3引起，尽管不排除钙蛋白酶-1的作用。
This study investigated the effects of elevated, physiological levels of intracellular free [Ca2+] on depolarization‐induced force responses, and on passive and active force production by the contractile apparatus in mechanically skinned fibres of toad iliofibularis muscle. Excitation–contraction (EC) coupling was retained after skinning and force responses could be elicited by depolarization of the transverse‐tubular (T‐) system. Raising the cytoplasmic [Ca2+] to ∼1 μm or above for 3 min caused an irreversible reduction in the depolarization‐induced force response by interrupting the coupling between the voltage sensors in the T‐system and the Ca2+ release channels in the sarcoplasmic reticulum. This uncoupling showed a steep [Ca2+] dependency, with 50% uncoupling at ∼1.9 μm Ca2+. The uncoupling occurring with 2 μm Ca2+ was largely prevented by the calpain inhibitor leupeptin (1 mm). Raising the cytoplasmic [Ca2+] above 1 μm also caused an irreversible decline in passive force production in stretched skinned fibres in a manner graded by [Ca2+], though at a much slower relative rate than loss of coupling. The progressive loss of passive force could be rapidly stopped by lowering [Ca2+] to 10 nm, and was almost completely inhibited by 1 mm leupeptin but not by 10 μm calpastatin. Muscle homogenates preactivated by Ca2+ exposure also evidently contained a diffusible factor that caused damage to passive force production in a Ca2+‐dependent manner. Western blotting showed that: (a) calpain‐3 was present in the skinned fibres and was activated by the Ca2+exposure, and (b) the Ca2+ exposure in stretched skinned fibres resulted in proteolysis of titin. We conclude that the disruption of EC coupling occurring at elevated levels of [Ca2+] is likely to be caused at least in part by Ca2+‐activated proteases, most likely by calpain‐3, though a role of calpain‐1 is not excluded.
反应性二硫化物通过氧化反应触发肌浆网中的 Ca2 释放。
DOI：
--
发表时间：
1989
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Zaidi,NF;Lagenaur,CF;Abramson,JJ;Pessah,I;Salama,G
通讯作者：
Salama,G