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Molecular analysis of ion transporters in plant and cyanobacteria involved in response to high osmolality

植物和蓝藻中参与高渗透压反应的离子转运蛋白的分子分析

基本信息

批准号：
17380064
负责人：
UOZUMI Nobuyuki
金额：
$ 6.46万
依托单位：
Tohoku University (2006)Nagoya University (2005)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17380064/
关键词：
osmolality tonoplast HKT positive residue Arabidopsis salt cyanobacteria pH sensor KCO系液泡膜 Na Hアンチポーター AtHKT1 Ktr カリウムナトリウム

项目摘要

1. We have identified the localization of Arabidopsis Na transporter AtHKT1 in plant cells. The specificity of the anti-AtHKT1 antibodies was confirmed. Immunoelectron microscopy using the antibodies demonstrate that AtHKT1 is targeted to the plasma membrane in xylem parenchyma cells in leaves.2. AtHKT1 disruption alleles caused large increases in the Na content of the xylem sap and conversely reduced the Na content of the phloem sap. The athkt1 mutant alleles had a smaller and inverse influence on the K content compared with the Na content to the xylem, suggesting that K transport may be indirectly affected. The expression of AtHKT1 was modulated not only by the concentrations of Na and K but also by the osmolality of non-ionic compounds.3. The conserved Arg near the middle of the M2D segment was essential for the K^+ transport of KtrB. We propose the existence of the salt bridge(s) between the positive residues in the M2D and the conserved negative residues in the pore region, which reduces an electrostatic barrier against cation permeation caused by the positive residue(s) and that stabilizes the transporter configuration.4. Mutation of a conserved His-157 in the second pore loop of KtrB, drastically reduced the activity of the K^+ transporter KtrABE system from Synechocystis PCC 6803. The result suggests that His-157 plays an essential role in the K^+ transport activity of the transporter system.5. H transport activity of one of Synechocystis Na/H transporters was measured using the inverted membrane vesicle of E. coli expressing the protein. We have identified the charged residues essential for the K transport.

1.我们已经确定了拟南芥钠转运蛋白AtHKT 1在植物细胞中的定位。证实了抗AtHKT 1抗体的特异性。免疫电镜显示AtHKT 1定位于叶片木质部薄壁细胞的质膜上. AtHKT 1破坏等位基因引起木质部汁液的Na含量的大幅度增加，相反地降低了韧皮部汁液的Na含量。athkt 1突变体等位基因有一个较小的和相反的影响K含量相比，Na含量的木质部，这表明K运输可能会间接影响。AtHKT 1的表达不仅受Na和K浓度的调节，还受非离子化合物渗透压的调节. M2 D片段中部附近的保守Arg对KtrB的K^+转运至关重要。我们提出在M2 D中的正性残基和孔区域中保守的负性残基之间存在盐桥，这降低了由正性残基引起的对阳离子渗透的静电屏障，并且稳定了转运体构型. KtrB第二孔环中保守His-157的突变大大降低了集胞藻PCC 6803的K^+转运蛋白KtrABE系统的活性。结果表明His-157在转运系统的K^+转运活性中起着重要作用.利用E.大肠杆菌表达该蛋白。我们已经确定了K运输所必需的带电残基。