Different effects of cardiac versus skeletal muscle regulatory proteins on in vitro measures of actin filament speed and force
Different effects of cardiac versus skeletal muscle regulatory proteins on in vitro measures of actin filament speed and force
复制标题
DOI：
10.1113/jphysiol.2005.084194
复制
发表时间：
2005-08-01
影响因子：
5.5
通讯作者：
Regnier, M
中科院分区：
文献类型：
--
作者：
Clemmens, EW;Entezari, M;Regnier, M
哺乳动物心肌和骨骼肌表达细丝调节蛋白的独特亚型，肌钙蛋白（Tn）和原肌球蛋白（Tm），这些不同亚型在细丝调节中的意义尚未明确。研究横纹肌细丝调节机制的体外和皮肤细胞研究经常使用Tn、Tm和肌球蛋白异质混合物，报告结果的差异可能是由这些蛋白质的不同组合解释的。在这里，我们使用体外运动和力（微针）测定来研究心脏和骨骼的Tn和Tin亚型对肌动蛋白重肌凝蛋白（HMM）力学的影响。当与骨骼HMM相互作用时，由心脏Tn/Tm或骨骼Tn/Tm重建的细丝表现出相似的速度-钙关系，并且在pCa 5（与未调节的肌动蛋白丝相比）显著增加了最大速度和每丝长度的力（F/l）。然而，骨骼调节蛋白对F/l的增加更大。相对于骨架Tn/Tm的细丝，骨架Tn和心脏Tin异质组合的细丝重构降低了所有[Ca2+]的滑动速度。最后,对于丝重组与任何异构Tn和Tm亚型,力不强,不受监管的肌动蛋白丝。综合结果表明：(1)心脏调节蛋白限制了力的变构增强；(2)在研究Ca2+对过桥结合和动力学的调节以及心脏和骨骼肌之间的机制差异时，Tn和Tm异构体的同质性是重要的。
Mammalian cardiac and skeletal muscle express unique isoforms of the thin filament regulatory proteins, troponin (Tn) and tropomyosin (Tm), and the significance of these different isoforms in thin filament regulation has not been clearly identified. Both in vitro and skinned cellular studies investigating the mechanism of thin filament regulation in striated muscle have often used heterogeneous mixtures of Tn, Tm and myosin isoforms, and variability in reported results might be explained by different combinations of these proteins. Here we used in vitro motility and force (microneedle) assays to investigate the influence of cardiac versus skeletal Tn and Tin isoforms on actin-heavy meromyosin (HMM) mechanics. When interacting with skeletal HMM, thin filaments reconstituted with cardiac Tn/Tm or skeletal Tn/Tm exhibited similar speed-calcium relationships and significantly increased maximum speed and force per filament length (F/l) at pCa 5 (versus unregulated actin filaments). However, augmentation of F/l was greater with skeletal regulatory proteins. Reconstitution of thin filaments with the heterogeneous combination of skeletal Tn and cardiac Tin decreased sliding speeds at all [Ca2+] relative to thin filaments with skeletal Tn/Tm. Finally, for filaments reconstituted with any heterogeneous mix of Tn and Tm isoforms, force was not potentiated over that of unregulated actin filaments. Combined the results suggest (1) that cardiac regulatory proteins limit the allosteric enhancement of force, and (2) that Tn and Tm isoform homogeneity is important when studying Ca2+ regulation of crossbridge binding and kinetics as well as mechanistic differences between cardiac and skeletal muscle.