Vasekina A V, Yershov P V, Reshetova O S, Tikhonova T V, Lunin V G, Trofimova M S, Babakov A V
All-Russia Institute of Agricultural Biotechnology, Russian Academy of Agricultural Sciences, Moscow 127550, Russia.
Biochemistry (Mosc). 2005 Jan;70(1):100-7. doi: 10.1007/s10541-005-0057-8.
One of the protective mechanisms used by plants to survive under conditions of salt stress caused by high NaCl concentration is the removal of Na+ from the cytoplasm. This mechanism involves a number of Na+/H+-antiporter proteins that are localized in plant plasma and vacuolar membranes. Due to the driving force of the electrochemical H+ gradient created by membrane H+-pumps (H+-ATPases and vacuolar H+-pyrophosphatases), Na+/H+-antiporters extrude sodium ions from the cytoplasm in exchange for protons. In this study, we have identified the gene for the barley vacuolar Na+/H+-antiporter HvNHX2 using the RACE (rapid amplification of cDNA ends)-PCR (polymerase chain reaction) technique. It is shown that the identified gene is expressed in roots, stems, and leaves of barley seedlings and that it presumably encodes a 59.6 kD protein composed of 546 amino acid residues. Antibodies against the C-terminal fragment of HvNHX2 were generated. It is shown that the quantity of HvNHX2 in tonoplast vesicles isolated from roots of barley seedlings remains the same, whereas the rate of Na+/H+ exchange across these membranes increases in response to salt stress. The 14-3-3-binding motif Lys-Lys-Glu-Ser-His-Pro (371-376) was detected in the HvNHX2 amino acid sequence, which is suggestive of possible involvement of the 14-3-3 proteins in the regulation of HvNHX2 function.
植物在高NaCl浓度引起的盐胁迫条件下生存所采用的保护机制之一是将Na⁺从细胞质中去除。该机制涉及一些位于植物质膜和液泡膜上的Na⁺/H⁺反向转运蛋白。由于膜H⁺泵（H⁺-ATP酶和液泡H⁺-焦磷酸酶）产生的电化学H⁺梯度的驱动力，Na⁺/H⁺反向转运蛋白将钠离子从细胞质中挤出以交换质子。在本研究中，我们使用RACE（cDNA末端快速扩增）-PCR（聚合酶链反应）技术鉴定了大麦液泡Na⁺/H⁺反向转运蛋白HvNHX2的基因。结果表明，鉴定出的基因在大麦幼苗的根、茎和叶中表达，并且推测它编码一种由546个氨基酸残基组成的59.6 kD蛋白质。产生了针对HvNHX2 C末端片段的抗体。结果表明，从大麦幼苗根部分离的液泡膜小泡中HvNHX2的量保持不变，而跨这些膜的Na⁺/H⁺交换速率响应盐胁迫而增加。在HvNHX2氨基酸序列中检测到14-3-3结合基序Lys-Lys-Glu-Ser-His-Pro（371-376），这表明14-3-3蛋白可能参与HvNHX2功能的调节。
