McElhinny A S, Kolmerer B, Fowler V M, Labeit S, Gregorio C C
Department of Cell Biology and Anatomy, University of Arizona, Tucson, Arizona 85724, USA.
J Biol Chem. 2001 Jan 5;276(1):583-92. doi: 10.1074/jbc.M005693200.
Strict regulation of actin thin filament length is critical for the proper functioning of sarcomeres, the basic contractile units of myofibrils. It has been hypothesized that a molecular template works with actin filament capping proteins to regulate thin filament lengths. Nebulin is a giant protein ( approximately 800 kDa) in skeletal muscle that has been proposed to act as a molecular ruler to specify the thin filament lengths characteristic of different muscles. Tropomodulin (Tmod), a pointed end thin filament capping protein, has been shown to maintain the final length of the thin filaments. Immunofluorescence microscopy revealed that the N-terminal end of nebulin colocalizes with Tmod at the pointed ends of thin filaments. The three extreme N-terminal modules (M1-M2-M3) of nebulin bind specifically to Tmod as demonstrated by blot overlay, bead binding, and solid phase binding assays. These data demonstrate that the N terminus of the nebulin molecule extends to the extreme end of the thin filament and also establish a novel biochemical function for this end. Two Tmod isoforms, erythrocyte Tmod (E-Tmod), expressed in embryonic and slow skeletal muscle, and skeletal Tmod (Sk-Tmod), expressed late in fast skeletal muscle differentiation, bind on overlapping sites to recombinant N-terminal nebulin fragments. Sk-Tmod binds nebulin with higher affinity than E-Tmod does, suggesting that the Tmod/nebulin interaction exhibits isoform specificity. These data provide evidence that Tmod and nebulin may work together as a linked mechanism to control thin filament lengths in skeletal muscle.
严格调控肌动蛋白细肌丝长度对于肌节（肌原纤维的基本收缩单位）的正常功能至关重要。据推测，一种分子模板与肌动蛋白丝封端蛋白共同作用来调控细肌丝长度。伴肌动蛋白是骨骼肌中的一种巨型蛋白质（约800 kDa），有人提出它可作为分子尺来确定不同肌肉特有的细肌丝长度。原肌球蛋白（Tmod）是一种位于细肌丝尖端的封端蛋白，已被证明可维持细肌丝的最终长度。免疫荧光显微镜检查显示，伴肌动蛋白的N末端与原肌球蛋白在细肌丝的尖端共定位。通过印迹覆盖、磁珠结合和固相结合试验证明，伴肌动蛋白的三个极端N末端模块（M1 - M2 - M3）与原肌球蛋白特异性结合。这些数据表明，伴肌动蛋白分子的N末端延伸至细肌丝的最末端，并且也为此末端确立了一种新的生化功能。两种原肌球蛋白异构体，即在胚胎和慢收缩骨骼肌中表达的红细胞原肌球蛋白（E - Tmod），以及在快收缩骨骼肌分化后期表达的骨骼肌原肌球蛋白（Sk - Tmod），在重叠位点与重组N末端伴肌动蛋白片段结合。Sk - Tmod与伴肌动蛋白的结合亲和力高于E - Tmod，这表明原肌球蛋白/伴肌动蛋白相互作用具有异构体特异性。这些数据提供了证据，表明原肌球蛋白和伴肌动蛋白可能作为一种连锁机制共同作用来控制骨骼肌中的细肌丝长度。
