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Effects of cold-induced dysfunction of vacuoles upon homeostasis in intracellular ionic environment.

寒冷诱导的液泡功能障碍对细胞内离子环境稳态的影响。

基本信息

批准号：
03454005
负责人：
YOSHIDA Shizuo
金额：
$ 4.16万
依托单位：
The Institute of Low Temperature Science, Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1992
项目状态：
已结题

项目摘要

Chilling sensitivity of mung bean cultured cells varied markedly depending on the growth phases. At the initial stages after subculture, cells were relatively insensitive to chilling, but chilling sensitivity increased abruptly thereafter and reached a maximum at the early stage of exponential growth. Upon further culturing, cell become chilling-tolerant, becoming most tolerant at the stage of stationary growth. We observed that cold sensitivity of the tonoplast H^+-ATPase in vivo changed markedly as a function of growth cycle. the enzyme was very sensitive to cold at the early stage of exponential growth and less sensitive at the late stage of exponential growth. Therefore, there exists an intimate correlation between sensitivities of cells to cold and the coldinsability of the H^+-ATPase. Biochemical studies on the cold-induced inactivation of the tonoplast H^+-ATPase indicated that a selective release of the peripheral subunits as a whole from the membrane sectors (16 kDa subunit).Changes in cytoplasmic pH in mung bean cultured cells were observed utilizing a cryomicroscope and a fluorescence ratio imaging system with special reference to the variations in chilling sensitivity during cell growth. In the extremely chilling-sensitive cells at the early stage of exponential growth, cytoplasmic pH was reduced abruptly upon cold incubation at 0ﾟC. However, in chilling- tolerant cells at the late stage of exponential growth, no change in the cytoplasmic pH was detected even after protracted chilling for up to 72 h.From the results, it is suggested that vacuolar proton translocating systems has an important role for maintaining of cell homeostasis, especially under such a cold environment.

绿豆培养细胞的冷敏感性随生长阶段的不同而有明显的差异。继代培养初期，细胞对低温相对不敏感，但此后低温敏感性急剧增加，并在指数生长初期达到最大值。在进一步培养时，细胞变得耐冷，在稳定生长阶段变得最耐冷。我们观察到，液泡膜H^+-ATPase的冷敏感性在体内作为生长周期的函数发生显著变化。该酶在指数生长初期对低温非常敏感，在指数生长后期对低温不敏感。因此，细胞对冷的敏感性与H^+-ATP酶的耐冷性之间存在着密切的关系。低温诱导液泡膜H^+-ATP酶失活的生化研究表明，液泡膜H^+-ATP酶的周边亚基（16 kDa亚基）整体选择性地从膜上释放出来。利用低温显微镜和荧光比率成像系统观察了绿豆培养细胞的细胞质pH变化，特别是细胞生长过程中对低温敏感性的变化。在极冷敏感细胞的指数生长的早期阶段，细胞质的pH值突然下降后，在0 ℃冷孵育。然而，在耐冷细胞的后期阶段的指数增长，没有变化的细胞质的pH值检测，即使在延长冷却长达72小时。从结果表明，液泡质子转运系统具有重要的作用，维持细胞内稳态，特别是在这样的寒冷环境。