STUDIES ON THE ROLE OF VACUOLES IN ION METABOLISMS IN PLANT.

在植物细胞无机离子代谢中，液泡是最重要的细胞器之一。液泡通常作为离子库和缓冲液来维持细胞质离子的稳态。对于这些功能，液泡膜离子转运系统起着重要的作用。本课题从生理学到分子生物学的角度，全面分析了植物细胞液泡在无机离子代谢中的作用。大麦液泡对磷的吸收活性不高。因此，我们试图测定悬浮培养的长春花细胞的磷吸收活性。已知这些细胞在质膜和液泡膜中都具有高的Pi摄取活性。与大麦一样，长春花液泡显示ATP和PPi依赖的Pi吸收。Pi吸收线性增加，直到20 mM的液泡外Pi。巴弗洛霉素（一种液泡膜H^+-ATPase的特异性抑制剂）也能抑制质膜对Pi的吸收。这些数据表明，Pi可能是在膜电位的驱动下通过液泡膜转运的。由于液泡膜H^+-ATPase是一个重要的影响因素，我们进一步分析了液泡膜H^+-ATPase的氧化还原调控，并利用离体轮藻节间细胞测定了Pi跨质膜转运网络与液泡膜的关系。

In inorganic ion metabolisms in plant cells, vacuole is one of the most important organelles. The vacuole is usually working as ion reservoir and buffer f or the cytoplasmic ion homeostasis. For these functions, the tonoplast ion transport systems are playing important roles. In the present project, we tried to totally analyze the role of the vacuole in inorganic ion metabolisms in plant cells by means of physiology to molecular biology.Last year, we have measured Pi uptake of isolated barley mesophyll vacuoles. Unfortunately, Pi uptake activity of barley vacuole is not high. Thus, we have tried to measure the Pi uptake activity of suspension cultured cells of Catharanthus roseus. These cells are known to have high activities of Pi uptake in both the plasma membrane and the tonoplast. As well as barley, Catharanthus vacuoles showed both ATP and PPi dependent Pi uptake. Pi uptake lineally increased until 20 mM extravacuolar Pi. Bafilomycin ; a specific inhibitor of the tonoplast H^+-ATPase also inhibited Pi uptake across the tonoplast. All these data suggested that Pi might be transported across the tonoplast driven by the membrane potential. Since the tonoplast H^+-ATPase is an important factor, we further analyzed the redox control of the tonoplast H^+-ATPase.We are also measuring the relationship between Pi transport network across the plasma membrane and the tonoplst using isolated Chara internodal cells.