Stac3 has a direct role in skeletal muscle-type excitation-contraction coupling that is disrupted by a myopathy-causing mutation
Stac3 has a direct role in skeletal muscle-type excitation-contraction coupling that is disrupted by a myopathy-causing mutation
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DOI：
10.1073/pnas.1612441113
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发表时间：
2016-09-27
影响因子：
11.1
通讯作者：
Beam, Kurt G.
中科院分区：
文献类型：
--
作者：
Polster, Alexander;Nelson, Benjamin R.;Beam, Kurt G.
在骨骼肌中，质膜中的Ca（v）1.1和肌浆网（SR）中的1型兰尼碱受体（RyR 1）之间的构象偶联被认为是兴奋-收缩（EC）偶联SR释放Ca 2+和逆行偶联（RyR 1通过Ca（v）1.1增加Ca 2+电流的幅度）的基础。最近的研究表明，EC偶联失败，在肌肉从小鼠和鱼无效的蛋白Stac 3（SH 3和半胱氨酸丰富的结构域3），但没有建立的功能作用的Stac 3的Ca（v）1.1-RyR 1的相互作用。我们使用tsA 201细胞和Stac 3 KO肌管研究了这一点。虽然在tsA 201细胞中证实Stac 3可以支持Ca（v）1.1的表面表达，（与其辅助性β（1a）和α（2）-δ（1）亚基共表达）和产生大的Ca 2+电流，我们发现没有Stac 3，辅助性γ（1）亚基也支持Ca（v）1.1/β（1a）/α（2）-δ（1）的膜表达，但这种结合只产生微小的Ca 2+电流。在Stac 3 KO肌管中，质膜中的Ca（v）1.1减少，但仍很大。然而，Stac 3 KO肌管中剩余的Ca（v）1.1不产生可感知的Ca 2+电流或EC偶联Ca 2+释放。WT Stac 3在Stac 3 KO肌管中的表达完全恢复了Ca 2+电流和EC偶联Ca 2+释放，而Stac 3（W280 S）（含有美洲土著肌病突变）的表达部分恢复了Ca 2+电流，但仅略微恢复了EC偶联。我们的结论是，Ca（v）1.1的膜运输是便利的，但不需要，Stac 3，和Stac 3直接参与Ca（v）1.1和RyR 1之间的构象耦合。
In skeletal muscle, conformational coupling between Ca(v)1.1 in the plasma membrane and type 1 ryanodine receptor (RyR1) in the sarcoplasmic reticulum (SR) is thought to underlie both excitation-contraction (EC) coupling Ca2+ release from the SR and retrograde coupling by which RyR1 increases the magnitude of the Ca2+ current via Ca(v)1.1. Recent work has shown that EC coupling fails in muscle from mice and fish null for the protein Stac3 (SH3 and cysteine-rich domain 3) but did not establish the functional role of Stac3 in the Ca(v)1.1-RyR1 interaction. We investigated this using both tsA201 cells and Stac3 KO myotubes. While confirming in tsA201 cells that Stac3 could support surface expression of Ca(v)1.1 (coexpressed with its auxiliary beta(1a) and alpha(2)-delta(1) subunits) and the generation of large Ca2+ currents, we found that without Stac3 the auxiliary gamma(1) subunit also supported membrane expression of Ca(v)1.1/beta(1a)/alpha(2)-delta(1), but that this combination generated only tiny Ca2+ currents. In Stac3 KO myotubes, there was reduced, but still substantial Ca(v)1.1 in the plasma membrane. However, the Ca(v)1.1 remaining in Stac3 KO myotubes did not generate appreciable Ca2+ currents or EC coupling Ca2+ release. Expression of WT Stac3 in Stac3 KO myotubes fully restored Ca2+ currents and EC coupling Ca2+ release, whereas expression of Stac3(W280S) (containing the Native American myopathy mutation) partially restored Ca2+ currents but only marginally restored EC coupling. We conclude that membrane trafficking of Ca(v)1.1 is facilitated by, but does not require, Stac3, and that Stac3 is directly involved in conformational coupling between Ca(v)1.1 and RyR1.