Knockouts of Physcomitrella patens CHX1 and CHX2 Transporters Reveal High Complexity of Potassium Homeostasis
Knockouts of Physcomitrella patens CHX1 and CHX2 Transporters Reveal High Complexity of Potassium Homeostasis
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DOI：
10.1093/pcp/pct096
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发表时间：
2013-09-01
影响因子：
4.9
通讯作者：
Haro, Rosario
中科院分区：
文献类型：
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作者：
Mottaleb, Shady A.;Rodriguez-Navarro, Alonso;Haro, Rosario
本研究旨在利用模式植物小壶菌（Physcomitrella patens）来增加我们对CHX转运体在植物细胞中的功能的了解，其中已经鉴定出四个CHX基因PpCHX1-PpCHX4。其中两个基因PpCHX1和PpCHX2的表达水平与PpACT5基因大致相同，但另外两个基因的表达水平极低。PpCHX1和PpCHX2恢复了大肠杆菌突变体在低含K+培养基上的生长，表明它们介导的K+摄取可能是通过与H+的同调而激活的。相反，这些基因抑制了酿酒酵母中与kha1突变相关的缺陷，这表明它们可能介导K+/H+反港。ppchx1 -绿色荧光蛋白（GFP）融合蛋白在与高尔基标记共定位的葡萄原生质体中的瞬时表达。在类似的实验中，PpCHX2-GFP蛋白似乎定位在细胞质和质膜上。我们构建了Delta Ppchx1和Delta Ppchx2单突变系，以及Delta Ppchx2 Delta Pphak1双突变系。在所有测试条件下，单突变体植株生长正常，K+和Rb+含量正常；Delta Ppchx2突变没有增加Delta Pphak1植株的缺陷。在长期实验中，Delta Ppchx2植株的Rb+保留率略高于野生型植株，这表明Ppchx2介导Rb+从液泡转移到细胞质或从细胞质转移到外部介质，与其他转运体平行。从技术上讲，很难区分这两种可能性。我们认为几个家族的K+转运体通过介导K+/H+反转运或K+-H+同调参与细胞器的pH稳态。
This study aims to increase our understanding of the functions of CHX transporters in plant cells using the model plant Physcomitrella patens, in which four CHX genes have been identified, PpCHX1-PpCHX4. Two of these genes, PpCHX1 and PpCHX2, are expressed at approximately the same level as the PpACT5 gene, but the other two genes show an extremely low expression. PpCHX1 and PpCHX2 restored growth of Escherichia coli mutants on low K+-containing media, suggesting that they mediated K+ uptake that may be energized by symport with H+. In contrast, these genes suppressed the defect associated with the kha1 mutation in Saccharomyces cerevisiae, which suggests that they might mediate K+/H+ antiport. PpCHX1-green fluorescent protein (GFP) fusion protein transiently expressed in P. patens protoplasts co-localized with a Golgi marker. In similar experiments, the PpCHX2-GFP protein appeared to localize to tonoplast and plasma membrane. We constructed the Delta Ppchx1 and Delta Ppchx2 single mutant lines, and the Delta Ppchx2 Delta Pphak1 double mutant. Single mutant plants grew normally under all the conditions tested and exhibited normal K+ and Rb+ influxes; the Delta Ppchx2 mutation did not increase the defect of Delta Pphak1 plants. In long-term experiments, Delta Ppchx2 plants showed slightly higher Rb+ retention than wild-type plants, which suggests that PpCHX2 mediates the transfer of Rb+ either from the vacuole to the cytosol or from the cytosol to the external medium in parallel with other transporters. The distinction between these two possibilities is technically difficult. We suggest that K+ transporters of several families are involved in the pH homeostasis of organelles by mediating either K+/H+ antiport or K+-H+ symport.