Molecular biological studies of ryanodine receptor/calcium release channel

本研究的目的是在全动物水平上阐明ryanodine受体/钙释放通道的生理功能。因此，我们首先试图寻找或产生突变动物的ryanodine受体基因。由于在果蝇系统中有很多已知的突变体，我们分析了黑腹果蝇的一种ryanodine受体基因并进行了染色体定位（FEBS） Lett. 337,81)；然而，在染色体位置上没有已知的突变体。然后，我们尝试用胚胎干细胞制备缺乏骨骼肌ryanodine受体的突变小鼠。在对产生的突变小鼠的第一次分析中，我们发现骨骼肌ryanodine受体是肌肉成熟和兴奋-收缩耦合的重要分子（Nature 369,556）。电镜观察发现，在突变肌中横管膜和肌浆网膜之间有窄尺寸的三联结，这表明骨骼肌ryanodine受体决定了膜之间的大小，但对连接的产生不是必需的（proc . natl . acad . scia . usa .92,3391）。此外，我们在突变肌肉中发现了脑型ryanodine受体的表达，并揭示了其药理学特性，同时不能促进骨骼肌兴奋-收缩耦联（EMBO j .14,2999）。综上所述，我们通过基因敲除小鼠的产生和分析确定了骨骼肌ryanodine受体的生理功能，并显示了ryanodine受体shbtypes之间的差异作用。

The aim of this research is elucidating physiological functions of teh ryanodine receptor/calcium ralease channel in a whole animallevel. Thus, we first tried to find or generate mutant animals in the ryanodine receptor gene. Because so many mutants are known in fruit fly system, we analyzed a ryanodine receptor gene in Drosophila melanogaster and the chromosomal mapping was done (FEBS) Lett. 337,81) ; however, no mutant is known at the located chromosomal position. Then, we tried to prepare mutnat mice lacking the skeletal muscle ryanodine receptor using embyonic stem cells. In the first analysis of the generated mutant mice, we showed that the skeletal muscle ryanodine receptor is an essential molecule for muscle maturation and excitation-contraction coupling (Nature 369,556). An electron-microscopy found triad juctions with a narrow size between transverse-tuble and sarcoplasmi reticulum membranes in the mutnat muscle, indicationg that the skeletal muscle ryanodine receptor determines the size between the membranes but is not essential for generation of the junction (Proc.Natl.Acad.Sci.USA.92,3391). Moreover, we found expression of the brain-type of ryanodine receptor in mutant muscle, and revealedits pharmoacological properties accompanying inability to contribute skeletal muscle exicitation-conrtraction couplina (EMBO J.14,2999).In summary, we defined the phisiological function of the skeletal muscle ryanodine receptor by generation and analysis of knockout mice, and showed the differential role between ryanodine receptor shbtypes.