Four and a half LIM protein 1 binds myosin-binding protein C and regulates myosin filament formation and sarcomere assembly
Four and a half LIM protein 1 binds myosin-binding protein C and regulates myosin filament formation and sarcomere assembly
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DOI：
10.1074/jbc.m512552200
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发表时间：
2006-03-17
影响因子：
4.8
通讯作者：
Brown, S
中科院分区：
文献类型：
--
作者：
McGrath, MJ;Cottle, DL;Brown, S
4个半LIM蛋白1 （FHL1/SLIM1）在骨骼肌和心肌中高表达；然而，FHL1的功能尚不清楚。酵母双杂交筛选鉴定出慢型骨骼肌球蛋白结合蛋白C作为FHL1的结合伙伴。肌凝蛋白结合蛋白C是横纹肌中主要的肌凝蛋白相关蛋白，可增强肌凝蛋白粗丝的横向结合和稳定性，调节肌动球蛋白相互作用。重组蛋白和内源性蛋白的共免疫沉淀证实了FHL1与肌球蛋白结合蛋白C的相互作用。重组FHL2和FHL3也结合肌球蛋白结合蛋白C。FHL1破坏了肌球蛋白结合蛋白C与重组肌球蛋白丝的共沉淀，表明FHL1可能与肌球蛋白竞争肌球蛋白与肌球蛋白结合蛋白C的结合。在完整骨骼肌和分离的肌原纤维中，FHL1定位于i带、m线和肌膜，与肌球蛋白结合蛋白C在完整骨骼肌的肌膜共定位。此外，在分离的肌原纤维中，m线的FHL1染色似乎部分延伸到a带的c区，在那里它与肌球蛋白结合蛋白c共定位。在分化的C2C12细胞中，FHL1的过度表达诱导“囊状”肌管形成（myosac），与z线和肌球蛋白粗丝组装受损有关。这种表型通过肌球蛋白结合蛋白C的共同表达得以挽救。使用RNAi敲低FHL1导致肌球蛋白粗丝形成受损，并减少肌球蛋白结合蛋白C进入肌节的结合。本研究发现FHL1是肌球蛋白结合蛋白C活性的一种新的调节因子，并表明FHL1在肌节组装中的作用。
Four and a half LIM protein 1 (FHL1/SLIM1) is highly expressed in skeletal and cardiac muscle; however, the function of FHL1 remains unknown. Yeast two-hybrid screening identified slow type skeletal myosin-binding protein C as an FHL1 binding partner. Myosin-binding protein C is the major myosin-associated protein in striated muscle that enhances the lateral association and stabilization of myosin thick filaments and regulates actomyosin interactions. The interaction between FHL1 and myosin-binding protein C was confirmed using co-immunoprecipitation of recombinant and endogenous proteins. Recombinant FHL2 and FHL3 also bound myosin-binding protein C. FHL1 impaired co-sedimentation of myosin-binding protein C with reconstituted myosin filaments, suggesting FHL1 may compete with myosin for binding to myosin-binding protein C. In intact skeletal muscle and isolated myofibrils, FHL1 localized to the I-band, M-line, and sarcolemma, co-localizing with myosin-binding protein C at the sarcolemma in intact skeletal muscle. Furthermore, in isolated myofibrils FHL1 staining at the M-line appeared to extend partially into the C-zone of the A-band, where it co-localized with myosin-binding protein C. Overexpression of FHL1 in differentiating C2C12 cells induced "sac-like" myotube formation (myosac), associated with impaired Z-line and myosin thick filament assembly. This phenotype was rescued by co-expression of myosin-binding protein C. FHL1 knockdown using RNAi resulted in impaired myosin thick filament formation associated with reduced incorporation of myosin-binding protein C into the sarcomere. This study identified FHL1 as a novel regulator of myosin-binding protein C activity and indicates a role for FHL1 in sarcomere assembly.