Calpain 1-gamma filamin interaction in muscle cells: a possible in situ regulation by PKC-alpha.
Calpain 1-gamma filamin interaction in muscle cells: a possible in situ regulation by PKC-alpha.
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发表时间：
2006
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通讯作者：
F. Raynaud;Carole Jond-Necand;A. Marcilhac;D. Fürst;Y. Benyamin
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作者：
F. Raynaud;Carole Jond-Necand;A. Marcilhac;D. Fürst;Y. Benyamin
钙蛋白酶是钙依赖性半胱氨酸蛋白酶家族，参与细胞骨架重塑和肌肉分化。在最近的一项研究中，我们观察到钙蛋白酶1在肌肉收缩装置中的存在，特别是在N1和N2线。发现这种钙蛋白酶亚型通过Z盘和肋连接中关键蛋白的蛋白水解参与肌纤维的降解。这项研究的目的是确定γ-细丝蛋白-细丝蛋白家族的一种特定肌肉亚型-是否是钙蛋白酶1底物，并验证这种相互作用。γ-细丝蛋白是位于Z线和肌膜下的主要肌肉结构蛋白。这种蛋白质是将肌膜结合到肌节结构的链的组分。在这项研究中，我们发现，γ-细丝蛋白形成了一个稳定的复合物在体外和细胞中的钙蛋白酶1在没有钙刺激。我们还将结合结构域定位在γ-细丝蛋白的C-末端，在铰链2区的丝氨酸2626和丝氨酸2627之间具有切割位点。钙蛋白酶1的催化亚基（80 kDa）和调节亚基（28 kDa）均参与与γ-细丝蛋白的高亲和力结合。此外，我们发现，在COS细胞中，PKC α对细丝蛋白C-末端结构域的磷酸化保护γ-细丝蛋白免受钙蛋白酶1的蛋白水解。刺激肌管中的PKC活性，防止钙蛋白酶引起的γ-细丝蛋白水解，并导致肌管粘附增加。
Calpains are a family of calcium-dependent cysteine-proteases involved in cytoskeleton remodelling and muscle differentiation. In a recent study, we observed the presence of calpain 1 in the muscle contractile apparatus and specifically in the N1- and N2-lines. This calpain isoform was found to be involved in the degradation of muscle fibres via proteolysis of key proteins in Z-disk and costameric junctions. The goal of this study was to determine whether gamma-filamin--a specific muscle isoform of the filamin family--is a calpain 1 substrate and to characterise this interaction. Gamma-filamin is a major muscle architectural protein located in the Z-line and under the sarcolemmal membrane. This protein is a component of the chain binding the sarcolemma to the sarcomeric structure. In this study, we found that gamma-filamin formed a stable complex in vitro and in cells with calpain 1 in the absence of calcium stimulation. We also located the binding domains in the C-terminus of gamma-filamin with a cleavage site between serine 2626 and serine 2627 in the hinge 2 region. The catalytic (80 kDa) and regulatory (28 kDa) subunits of calpain 1 are both involved in high affinity binding at gamma-filamin. Moreover, we showed that phosphorylation of the filamin C-terminus domain by PKC alpha protected gamma-filamin against proteolysis by calpain 1 in COS cells. Stimulation of PKC activity in myotubes, prevented gamma-filamin proteolysis by calpain and resulted in an increase in myotube adhesion.