Functional analysis of the Na+/H+ antiporter encoding genes of the cyanobacterium Synechocystis PCC 6803
Functional analysis of the Na+/H+ antiporter encoding genes of the cyanobacterium Synechocystis PCC 6803
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DOI：
10.1023/a:1015281906254
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发表时间：
2002-04-01
影响因子：
2.8
通讯作者：
Hagemann, M
中科院分区：
文献类型：
--
作者：
Elanskaya, IV;Karandashova, IV;Hagemann, M
研究了由nhaS1（slr-1727）、nhaS3（sll0689）、nhaS4（slr1595）和nhaS5（slr0415）编码的Na +/H+反向转运蛋白在蓝细菌集胞藻PCC 6803盐胁迫响应和内部pH调节中的作用。利用插入子诱变法构建了Na~+/H~+反向转运蛋白基因的单、双、三联突变体，PCR分析表明nhaS_1、nhaS_4、nhaS_5基因突变完全分离，突变体中只含有相应基因的失活拷贝。nhaS3编码的Na +/H+逆向转运蛋白是集胞藻生存所必需的，因为没有得到完全分离的突变体。稳态细胞内钠浓度和Na +/H+逆向转运活性被发现是相同的野生型和所有的突变体。野生型和突变体在补充有0.68 M或0.85 M NaCl的液体培养基中以及在pH 7.0、8.0或9.0缓冲的培养基中培养期间的生长速率没有差异。研究了Na +/H+反向转运蛋白基因的表达。在高盐或不同pH值下生长的野生型或单突变细胞中，未发现任何Na +/H+逆向转运蛋白编码基因表达的诱导。然而，在细胞的双重和三重突变体适应高盐或碱性pH值的一些剩余的Na +/H+逆向转运蛋白编码基因显示诱导。这些结果可能表明，一些Na +/H+反向转运蛋白可以在胁迫条件下相互取代，在集胞藻细胞缺乏一个以上的反向转运蛋白的活性。
The role of putative Na+/H+ antiporters encoded by nhaS1 (slr-1727), nhaS3 (sll0689), nhaS4 (slr1595), and nhaS5 (slr0415) in salt stress response and internal pH regulation of the cyanobacterium Synechocystis PCC 6803 was investigated. For this purpose the mutants (single, double, and triple) impaired in genes coding for Na+/H+ antiporters were constructed using the method of interposon mutagenesis, PC R analyses of DNA demonstrated that mutations in nhaS1, nhaS4, and nhaS5 genes were segregated completely and the mutants contained only inactivated copies of the corresponding genes. Na+/H+ antiporter encoded by nhaS3 was essential for viability of Synechocystis since no completely segregated mutants were obtained. The steady-state intracellular sodium concentration and Na+/H+ antiporter activities were found to be the same in the wild type and all mutants. No differences were found in the growth rates of wild type and mutants during their cultivation in liquid media supplemented with 0.68 M or 0.85 M NaCl as well as in media buffered at pH 7.0, 8.0, or 9.0. The expression of genes coding for Na+/H+ antiporters was studied. No induction of any Na+/H+ antiporter encoding gene expression was found in wild type or single mutant cells grown under high salt or at different pH values. Nevertheless, in cells of double and triple mutants adapted to high salt or alkaline pH some of the remaining Na+/H+ antiporter encoding genes showed induction. These results might indicate that some of Na+/H+ antiporters can functionally replace each other under stress conditions in Synechocystis cells lacking the activity of more than one antiporter.