STUDY ON THE DEVELOPMENT OF MOLECULAR BIOLOGY AND PHYSIOLOGY OF SA CHANNELS

SA通道的分子生物学和生理学发展研究

• 批准号: 09044283
• 负责人: SOKABE Masahiro
• 金额: $ 2.37万
• 依托单位: NAGOYA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for international Scientific Research
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09044283/
• 关键词: SA channel; cell volume regulation; endothelial cells; stretch stimulation; AFM; mec; mid-1; リモデリング; 遺伝子クローニング; ブロッカー; チャネル遺伝子; 伸展刺激装置; チャネル解析法; 原子間力顕微鏡

Stretch activated (SA) channels are expressed in a variety of cells and, thought to have important roles in fundamental cell functions. However their physiological functions are not yet known and their molecular entity has not yet be identified except for that in e. coli. To resolve these problems is the most important issue on the SA channel. The aim of this research project is to find out a clue to this goal through a collaboration between Japan and the U.S.As for the first problem, a cation selective SA channel has been found to have an important role in the regulatory volume decrease in A6 cells throuigh the estimation of whole cell activity of the SA channel by measuring intracellular Ca2+ incresae. Moreover the role of a cation selective SA channel has been clarified in stretch -induced cell remodeling in endothelial cells (Japan side). On the other hand ; the US researchers has developed a method to apply quantitative mechanical stimuli to cultured cells using canti-lever of AFM (atomic force microscope). They successfully explained the stretch induced-activity of heart cells by this method combined with sinultaneous whole cell recordings of SA channel currents.The second problem has been a tough one. We tried to clone a Drosophila gene homologous to mec gene from c.elegance, a putative SA channel gene, but failed. The US side tried to purify a SA channel-specific blocker from spider venoms, but yet to be done. However, at the last moment, we (Japan side) could identify a gene (mid-1) encoding cation selective SA channel from yeast. This is the first identified gene for the cation selective SA channel. By this finding we can evisage a variety of intriguing research projects including structure-function study. Based on this result, we will pursue further collaboration between Japan and the US.

拉伸激活（SA）通道在多种细胞中表达，被认为在基本细胞功能中起重要作用。然而，它们的生理功能尚不清楚，除了在大肠杆菌中，它们的分子实体尚未确定。解决这些问题是SA通道上最重要的问题。本研究项目的目的是通过日本和美国的合作来寻找实现这一目标的线索。对于第一个问题，通过测量细胞内Ca2+的增加来估计SA通道的全细胞活性，发现阳离子选择性SA通道在A6细胞的调节体积减少中起重要作用。此外，阳离子选择性SA通道在内皮细胞拉伸诱导的细胞重塑中的作用已被阐明（日本方面）。另一方面；美国研究人员开发了一种利用原子力显微镜（AFM）悬臂杠杆对培养细胞施加定量机械刺激的方法。他们成功地解释了心脏细胞的拉伸诱导活动，通过这种方法结合同时全细胞记录的SA通道电流。第二个问题很棘手。我们试图克隆一个与果蝇mec基因同源的果蝇SA通道基因，但没有成功。美方曾试图从蜘蛛毒液中纯化SA通道特异性阻滞剂，但尚未成功。然而，在最后一刻，我们（日本方面）从酵母中鉴定出一个编码阳离子选择性SA通道的基因（mid-1）。这是第一个确定的阳离子选择性SA通道基因。通过这一发现，我们可以设想各种有趣的研究项目，包括结构-功能研究。基于这一结果，我们将寻求日本和美国之间的进一步合作。

项目成果

Mosbacher J,Langer M,Horber JK,Sachs F.: "Voltage-dependent membrane displacements measured by atomic force microscopy." J.Gen Physiol.111 (1). 65-74 (1998)

Mosbacher J、Langer M、Horber JK、Sachs F.：“通过原子力显微镜测量电压依赖性膜位移。”

Mosbacher J, Langer M, Horber JK, Sachs F.: "Voltage-dependent membrane displacements measured by atomic force microscopy." J Gen Physiol.111(1). 65-74 (1998)

Mosbacher J、Langer M、Horber JK、Sachs F.：“通过原子力显微镜测量电压依赖性膜位移。”

Yu, W., Sokabe, M.: "Hypotonically induced whole cell currents in A6 cells : elationship with cell volume and cytoplasmic Ca^<2+>." Jpn J Physiol. 47 (6). 553-565 (1997)

Yu, W., Sokabe, M.：“A6 细胞中低渗诱导的全细胞电流：与细胞体积和细胞质 Ca^2 的关系。”

Naruse, K.Sai, X., Yokoyama, N., Sokabe,M.: "Uniaxialcyclic stretch indeces c-src activation and traslocation in human endothelial cells via SA channel activation." FEBS Lett.441. 111-115 (1998)

Naruse, K.Sai, X.、Yokoyama, N.、Sokabe,M.：“单轴循环拉伸通过 SA 通道激活人内皮细胞中的 c-src 激活和易位。”

Akinlaja J,Sachs F.: "The breakdown of cell membranes by electrical and mechanical stress." Biophys J.75 (1). 247-54 (1998)

Akinlaja J，Sachs F.：“电应力和机械应力导致细胞膜破裂。”