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Molecular mechanisms of formation and maintenance of muscle contractile apparatus (myofibrils)

肌肉收缩装置（肌原纤维）形成和维持的分子机制

基本信息

批准号：
10304065
负责人：
OBINATA Takashi
金额：
$ 24.02万
依托单位：
CHIBA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2001
项目状态：
已结题

项目摘要

We examined molecular mechanisms of morphogenesis and maintenance of myofibrils by mainly focussing on cofilin (CF), an actin-binding protein, and C-protein, a myosin- and connectin-bindirig protein. The results are summarized as follow :1.Role of cofilin in myofibrillogenesis1) Excess of CF in muscle cells, which was caused by cDNA transfection or injection of recombinant protein led to disruption of actin filaments and partial disorganization of myofibrils ; 2) In CF-deficient muscle cells, which were generated in mice by gene-manipulation, myofibrillar structures were severely damaged and muscle degeneration was induced in the region where CF-deficient muscle cells were concentrated ; 3) Inactivation of CF by increasing PIP2 in muscle cells also led to insufficient myofibril assembly.Based on these results, we conclude that CF is important not only in myofibril formation but also in maintenance of normal muscle structure.2. Molecular structure and function of C-protein1) Molecular structures of mouse fast skeletal (F) and slow skeletal (S) C-proteins were determined by cDNA technology, and their expression and assembly were clarified and compared with that of cardiac isoform ; 2) Actin-binding domain was specified in N-terminal region of cardiac C-protein ; 3) A novel mouse cardiac C-protein was discovered, which slightly differ in the C-terminal myosin-binding and connectin-binding region and functionally weak. The expression of this variant is up-regulated in aged heart.3. Role of actin-myosin interaction in myofibrillogenesisWe found that BDM (2,3-butanedione 2-monoxime), an inhibitor for myosin ATPase, suppresses, myfibrillogenesis and leads to myofibril disruption and thereby suggested that actin-myosin interaction plays a critical role in the early process of myofibrillogenesis.

我们研究了肌原纤维的形态发生和维持的分子机制，主要集中在cofilin（CF），肌动蛋白结合蛋白，和C蛋白，肌球蛋白和连接蛋白结合蛋白。结果表明：1. cofilin在肌原纤维发生中的作用1）通过基因转染或注射重组蛋白引起肌细胞内CF过量，导致肌动蛋白丝断裂和部分肌原纤维结构紊乱，2）通过基因操作产生CF缺陷肌细胞，CF缺陷肌细胞集中区域的肌原纤维结构严重受损，导致肌肉变性; 3）增加肌细胞中PIP 2对CF的失活也会导致肌原纤维组装不足，基于这些结果，我们得出结论：CF不仅在肌原纤维的形成中起重要作用，而且在维持正常肌肉结构中也起重要作用. C蛋白的分子结构与功能1）利用cDNA技术测定了小鼠快骨骼（F）和慢骨骼（S）C蛋白的分子结构，阐明了它们的表达和组装，并与心脏C蛋白的亚型进行了比较：2）在心脏C蛋白的N端区域发现了肌动蛋白结合域; 3）发现了一种新的小鼠心肌C蛋白，其C端肌球蛋白结合区和连接蛋白结合区略有不同，功能较弱。该变异体在老年心脏中表达上调.肌动蛋白-肌球蛋白相互作用在肌原纤维形成中的作用我们发现，BDM（2，3-丁二酮2-单肟），一种肌球蛋白ATP酶的抑制剂，抑制肌原纤维形成，并导致肌原纤维的破坏，从而表明肌动蛋白-肌球蛋白相互作用在肌原纤维形成的早期过程中起着关键作用。