An Insertion Sequence Prepares Pseudomonas putida S12 for Severe Solvent Stress*
An Insertion Sequence Prepares Pseudomonas putida S12 for Severe Solvent Stress*
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DOI：
10.1074/jbc.m007687200
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发表时间：
2001-02
期刊：
影响因子：
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通讯作者：
J. Wery;B. Hidayat;J. Kieboom;J. D. de Bont
中科院分区：
文献类型：
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作者：
J. Wery;B. Hidayat;J. Kieboom;J. D. de Bont
新的插入序列iss12在恶臭假单胞菌S12对突发性甲苯胁迫的耐受性中起关键作用。在正常培养条件下，p。putida S12基因组包含7个ISS12拷贝。然而，突然加入一个单独的阶段的甲苯后，生长到高细胞密度的p.p putida S12群体携带8个拷贝。该变异恶臭p.s 12群体的细胞存活率比“正常”恶臭p.s 12群体高1000倍。对额外的ISS12插入侧的核苷酸序列分析显示整合到srpS基因中。srps2与srpR形成一个基因簇，两者都被认为是溶剂抗性泵SrpABC的调节因子。SrpABC在恶臭p.p . putida S12耐溶剂性中起主要作用，是由甲苯诱导的。srp启动子活性的基础水平是野生型恶臭假单胞菌S12的7倍。在变体中引入完整的srpRS基因簇导致甲苯休克后存活频率急剧下降。这些发现有力地表明，ISS12对srpS的中断上调了溶剂泵的表达，使细菌能够耐受突然暴露于致死浓度的有毒溶剂。我们提出p.p putida S12利用ISS12作为突变元件产生多种突变，以迅速适应严峻的不利条件。
The novel insertion sequence ISS12plays a key role in the tolerance of Pseudomonas putida S12 to sudden toluene stress. Under normal culturing conditions theP. putida S12 genome contained seven copies of ISS12. However, a P. putida S12 population growing to high cell density after sudden addition of a separate phase of toluene carried eight copies. The survival frequency of cells in this variant P. putida S12 population was 1000 times higher than in “normal” P. putida S12 populations. Analysis of the nucleotide sequence flanking the extra ISS12 insertion revealed integration into the srpS gene. srpSforms a gene cluster with srpR and both are putative regulators of the solvent resistance pump SrpABC. SrpABC makes a major contribution to solvent tolerance in P. putida S12 and is induced by toluene. The basal level of srp promoter activity in the P. putida S12 variant was seven times higher than in wild-type P. putida S12. Introduction of the intact srpRS gene cluster in the variant resulted in a dramatic decrease of survival frequency after a toluene shock. These findings strongly suggest that interruption of srpS by ISS12 up-regulates expression of the solvent pump, enabling the bacterium to tolerate sudden exposure to lethal concentrations of toxic solvents. We propose that P. putida S12 employs ISS12 as a mutator element to generate diverse mutations to swiftly adapt when confronted with severe adverse conditions.