Proteomic Analyses Reveal the Mechanism of Dunaliella salina Ds-26-16 Gene Enhancing Salt Tolerance in Escherichia coli.
Proteomic Analyses Reveal the Mechanism of Dunaliella salina Ds-26-16 Gene Enhancing Salt Tolerance in Escherichia coli.
复制标题
蛋白质组学分析揭示杜氏盐藻 Ds-26-16 基因增强大肠杆菌耐盐性的机制
DOI：
10.1371/journal.pone.0153640
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发表时间：
2016
期刊：
影响因子：
3.7
通讯作者：
Chen D
中科院分区：
文献类型：
--
作者：
Wang Y;Hu B;Du S;Gao S;Chen X;Chen D
我们先前从4M盐胁迫下的杜氏盐藻(Dunaliella Salina)cDNA文库中筛选出了新的基因DS-26-16，发现该基因对广谱生物具有耐盐性，包括大肠杆菌、雨生红球藻和烟草。为了确定该基因的耐盐性机制，我们利用iTRAQ(相对和绝对定量等压标签)方法对过表达DS-26-16全长cDNA的大肠杆菌蛋白质组进行了研究。共鉴定出1610个蛋白质，占整个蛋白质组的39.4%。在559个差异蛋白质中，259个上调，300个下调。GO(基因本体)和KEGG(京都基因和基因组百科全书)浓缩分析鉴定了202个主要蛋白质，包括与氨基酸和有机酸代谢、能量代谢、碳代谢、ROS(活性氧物种)清除、膜蛋白和ABC(ATP结合盒)转运蛋白、肽聚糖合成以及5个上调转录因子有关的蛋白质。我们的iTRAQ数据表明，DS-26-16上调了大肠杆菌中的转录因子，通过渗透平衡、能量代谢和氧化应激保护来增强耐盐性。在过量表达DS-26-16开放阅读框架(ORF)的大肠杆菌中，也观察到了蛋白质组的变化。此外，pH、一氧化氮和甘油含量分析表明，DS-26-16的过表达增加了一氧化氮的含量，但对甘油含量没有影响，从而证实了DS-26-16通过降低胞间pH促进一氧化氮的合成是DS-26-16赋予大肠杆菌耐盐性的机制之一。
We previously screened the novel gene Ds-26-16 from a 4 M salt-stressed Dunaliella salina cDNA library and discovered that this gene conferred salt tolerance to broad-spectrum organisms, including E. coli (Escherichia coli), Haematococcus pluvialis and tobacco. To determine the mechanism of this gene conferring salt tolerance, we studied the proteome of E. coli overexpressing the full-length cDNA of Ds-26-16 using the iTRAQ (isobaric tags for relative and absolute quantification) approach. A total of 1,610 proteins were identified, which comprised 39.4% of the whole proteome. Of the 559 differential proteins, 259 were up-regulated and 300 were down-regulated. GO (gene ontology) and KEGG (Kyoto encyclopedia of genes and genomes) enrichment analyses identified 202 major proteins, including those involved in amino acid and organic acid metabolism, energy metabolism, carbon metabolism, ROS (reactive oxygen species) scavenging, membrane proteins and ABC (ATP binding cassette) transporters, and peptidoglycan synthesis, as well as 5 up-regulated transcription factors. Our iTRAQ data suggest that Ds-26-16 up-regulates the transcription factors in E. coli to enhance salt resistance through osmotic balance, energy metabolism, and oxidative stress protection. Changes in the proteome were also observed in E. coli overexpressing the ORF (open reading frame) of Ds-26-16. Furthermore, pH, nitric oxide and glycerol content analyses indicated that Ds-26-16 overexpression increases nitric oxide content but has no effect on glycerol content, thus confirming that enhanced nitric oxide synthesis via lower intercellular pH was one of the mechanisms by which Ds-26-16 confers salt tolerance to E. coli.