Mechanical stability of αT-catenin and its activation by force for vinculin binding.
Mechanical stability of αT-catenin and its activation by force for vinculin binding.
复制标题
αT-连环蛋白的机械稳定性及其通过强力激活纽蛋白结合。
DOI：
10.1091/mbc.e19-02-0102
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发表时间：
2019
影响因子：
3.3
通讯作者：
Yan,Jie
中科院分区：
文献类型：
--
作者：
Pang,SiMing;Le,Shimin;Kwiatkowski,AdamV;Yan,Jie
αT（睾丸）-连环蛋白是心脏中调节细胞-细胞粘附的关键因子，它直接将钙粘蛋白-连环蛋白复合物偶联到闰盘（ICD）的肌动蛋白细胞骨架上，闰盘是偶联心肌细胞的独特细胞-细胞连接。小鼠中α T-连环蛋白的缺失减少了斑块嗜蛋白2和连接蛋白43向ICD的募集。由于αT-catenin在心肌细胞肌动球蛋白收缩过程中受到机械拉伸，因此其活性可以通过机械力调节。为了深入了解力如何调节αT-catenin功能，我们研究了αT-catenin的假定力感应中间（M）结构域的机械稳定性，并确定了力如何影响黏着斑蛋白与αT-catenin的结合。我们发现：1）生理水平的力，<15 pN，足以展开三个M结构域; 2）含有黏着斑蛋白结合位点的M1结构域在106 pN时展开; 3）M1结构域的展开是高亲和力黏着斑蛋白结合所必需的。此外，我们定量了黏着斑蛋白与M1中机械暴露的结合位点的结合动力学和亲和力，并观察到α T-连环蛋白以低纳摩尔亲和力结合黏着斑蛋白。这些结果为αT-catenin的机械传感特性以及αT-catenin如何调节心肌细胞ICD处的细胞-细胞粘附提供了重要的新见解。
αT (Testes)-catenin, a critical factor regulating cell–cell adhesion in the heart, directly couples the cadherin-catenin complex to the actin cytoskeleton at the intercalated disk (ICD), a unique cell–cell junction that couples cardiomyocytes. Loss of αT-catenin in mice reduces plakophilin2 and connexin 43 recruitment to the ICD. Since αT-catenin is subjected to mechanical stretch during actomyosin contraction in cardiomyocytes, its activity could be regulated by mechanical force. To provide insight in how force regulates αT-catenin function, we investigated the mechanical stability of the putative, force-sensing middle (M) domain of αT-catenin and determined how force impacts vinculin binding to αT-catenin. We show that 1) physiological levels of force, <15 pN, are sufficient to unfold the three M domains; 2) the M1 domain that harbors the vinculin-binding site is unfolded at ∼6 pN; and 3) unfolding of the M1 domain is necessary for high-affinity vinculin binding. In addition, we quantified the binding kinetics and affinity of vinculin to the mechanically exposed binding site in M1 and observed that αT-catenin binds vinculin with low nanomolar affinity. These results provide important new insights into the mechanosensing properties of αT-catenin and how αT-catenin regulates cell–cell adhesion at the cardiomyocyte ICD.