Enhanced skeletal muscle contraction with myosin light chain phosphorylation by a calmodulin-sensing kinase
Enhanced skeletal muscle contraction with myosin light chain phosphorylation by a calmodulin-sensing kinase
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DOI：
10.1074/jbc.m702927200
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发表时间：
2007-07-13
影响因子：
4.8
通讯作者：
Stull, James T.
中科院分区：
文献类型：
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作者：
Ryder, Jeffrey W.;Lau, Kim S.;Stull, James T.
由于 Ca2+/钙调蛋白 (CaM) 依赖性骨骼肌肌球蛋白轻链激酶 (skMLCK) 导致肌球蛋白调节轻链 (RLC) 磷酸化，重复低频刺激会增强快肌骨骼肌的肌收缩力发展。我们培育了在骨骼肌中表达 skMLCK CaM 生物传感器的转基因小鼠，以确定 skMLCK 或 CaM 是否限制抽搐力增强。三个转基因小鼠品系在含有 IIa 型和 IIb 型纤维的快肌指长伸肌中 skMLCK 蛋白表达增加了 22 倍，而在含有 I 型和 IIa 型纤维的慢肌比目鱼肌中表达量相当。高表达系在低频电刺激下表现出更快速的 RLC 磷酸化和趾长伸肌的力量增强。令人惊讶的是，尽管快肌和慢肌 RLC 磷酸化均显着增强，但比目鱼肌中 skMLCK 的过度表达并未重现快肌增强反应。钙调蛋白与生物传感器结合的分析显示频率依赖性激活最大程度为 60%。由于 skMLCK 转基因表达比野生型激酶高 22 倍，因此 skMLCK 而不是钙调蛋白通常限制 RLC 磷酸化和抽搐力增强。激酶激活速率(10.6 s(-1))仅比收缩速率慢3.6倍，而失活速率(2.8 s(-1))比舒张速率慢12倍。体内激酶失活速度较慢且重复收缩，通过 RLC 磷酸化提供生化记忆。重要的是，RLC 磷酸化在 IIb 型快肌肌力增强中发挥着重要作用，但在 I 型或 IIa 型纤维中则不然。
Repetitive low frequency stimulation results in potentiation of twitch force development in fast-twitch skeletal muscle due to myosin regulatory light chain (RLC) phosphorylation by Ca2+/ calmodulin (CaM)-dependent skeletal muscle myosin light chain kinase (skMLCK). We generated transgenic mice that express an skMLCK CaM biosensor in skeletal muscle to determine whether skMLCK or CaM is limiting to twitch force potentiation. Three transgenic mouse lines exhibited up to 22-fold increases in skMLCK protein expression in fast-twitch extensor digitorum longus muscle containing type IIa and IIb fibers, with comparable expressions in slow-twitch soleus muscle containing type I and IIa fibers. The high expressing lines showed a more rapid RLC phosphorylation and force potentiation in extensor digitorum longus muscle with low frequency electrical stimulation. Surprisingly, overexpression of skMLCK in soleus muscle did not recapitulate the fast-twitch potentiation response despite marked enhancement of both fast-twitch and slow-twitch RLC phosphorylation. Analysis of calmodulin binding to the biosensor showed a frequency-dependent activation to a maximal extent of 60%. Because skMLCK transgene expression is 22-fold greater than the wild-type kinase, skMLCK rather than calmodulin is normally limiting for RLC phosphorylation and twitch force potentiation. The kinase activation rate (10.6 s(-1)) was only 3.6-fold slower than the contraction rate, whereas the inactivation rate (2.8 s(-1)) was 12-fold slower than relaxation. The slower rate of kinase inactivation in vivo with repetitive contractions provides a biochemical memory via RLC phosphorylation. Importantly, RLC phosphorylation plays a prominent role in skeletal muscle force potentiation of fast-twitch type IIb but not type I or IIa fibers.