Myosin: Structure, Function, Regulation and Disease
Myosin: Structure, Function, Regulation and Disease
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肌球蛋白：结构、功能、调节和疾病
DOI：
10.1096/fasebj.2020.34.s1.00126
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Peckham M
中科院分区：
文献类型：
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作者：
Peckham M
人肌球蛋白重链超家族包括12个不同的类别。2类是最大的，由14个肌球蛋白重链（MHC）基因（和1个假基因）组成。2类肌球蛋白重链形成两条重链的二聚体，并与2对轻链（必需和调节）结合。在13种2类肌球蛋白中，10种在横纹肌中表达，其余4种在非肌细胞和平滑肌中表达。所有的2类肌球蛋白形成细丝。在横纹肌中，肌丝精确地包含294个分子。相比之下，非肌肉肌球蛋白形成更短的细丝（约30个分子）。所有这些肌球蛋白的活性通过头部（包括马达结构域和杠杆）与卷曲螺旋的第一部分（亚片段2，S2）的相互作用来调节，以形成所谓的“相互作用头部”基序。非肌肉和平滑肌肌球蛋白头部还受到头部与尾部形成区（轻肌球蛋白，LMM）相互作用的调节，形成关闭分子。我们结合细胞生物学和结构方法，包括电子显微镜来了解2类肌球蛋白中S2和LMM致病突变的影响。纹状肌球蛋白（MyHCβ，MyHC 2a）的突变分别导致心脏和骨骼肌疾病，尽管MyHCβ远端LMM区域的突变主要与骨骼肌疾病相关。非肌肉肌球蛋白2A（NM 2A）突变可导致一系列出血性疾病。我们确定了特定的突变如何影响肌球蛋白形成肌丝的能力，以及它们如何影响其关闭状态的形成。例如，我们使用圆二色性来确定特定突变是否影响LMM的二级结构。我们使用负染色电子显微镜来评估LMM（与GST融合，以防止次晶体形成）的细丝形成，并且我们使用GFP标记的MHC来评估每种突变体掺入培养细胞、肌管和成年大鼠心肌细胞中的细丝的能力（帕克等人（2018）J. Mol. Biol. Biol）。我们最近解决了全长平滑肌肌球蛋白（SMM）在其关闭状态下的结构（Scarff et al.（2020）Nature）。这表明关闭状态是如何稳定的，使我们能够预测突变如何破坏它。再加上额外的结构研究，我们开始了解特定的突变如何影响肌球蛋白功能，以更好地了解疾病的过程。支持或资助信息本研究由医学研究理事会支持，MR/R 009406/1和MR/S 023593/1 MP。
The human myosin heavy chain superfamily comprises 12 different classes. Class 2 is the largest and is made up of 14 myosin heavy chain (MHC) genes (and 1 pseudogene). Class 2 myosin heavy chains form a dimer of two heavy chains, and associate with 2 pairs of light chains (essential and regulatory). Of the 13 types of class 2 myosins, 10 are expressed in striated muscle and the remaining 4 in non‐muscle cells and smooth muscle. All of the class 2 myosins form filaments. In striated muscle, the filaments contain precisely 294 molecules. In contrast, non‐muscle myosins form much shorter filaments (about 30 molecules). The activity of all of these myosins is regulated by an interaction of the heads (comprising the motor domain and lever) with the first part of the coiled coil (subfragment 2, S2) to form the so‐called ‘interacting heads’ motif. Non‐muscle and smooth muscle myosin heads are additionally regulated by an interaction of the head with the tail forming region (light meromyosin, LMM) to form a shutdown molecule.We use a combination of cell biology, and structural approaches, including electron microscopy to understand the effects of disease‐causing mutations in S2 and in LMM in class 2 myosins. Mutations in striated myosins (MyHCβ, MyHC2a) cause cardiac and skeletal muscle diseases respectively, although mutations in the distal LMM region of MyHCβ are predominantly associated with skeletal muscle disease. Mutations in non‐muscle myosin 2A (NM2A) cause a range of bleeding disorders. We determine how specific mutations affect the myosin's ability to form filaments and how they affect formation of its shutdown state. For example, we use circular dichroism to determine if specific mutations affect the secondary structure of LMM. We use negative stain electron microscopy to evaluate filament formation by the LMM (fused to GST, to prevent paracrystal formation) and we use GFP‐tagged MHC to evaluate the ability of each mutant to incorporate into filaments in cultured cells, myotubes and adult rat cardiomyocytes (Parker et al. (2018) J. Mol. Biol). We recently solved the structure of full‐length smooth muscle myosin (SMM) in its shutdown state (Scarff et al.. (2020) Nature) using CryoEM. This shows how the shutdown state is stabilised and enables us to predict how mutations might disrupt it. Together with additional structural studies, we are starting to understand how specific mutations affect myosin function, to gain a better understanding of the disease process.Support or Funding InformationThis research is supported by the Medical Research Council, MR/R009406/1 and MR/S023593/1 to MP.