TnpR Encoded by an ISPpu12 Isoform Regulates Transposition of Two Different ISL3-Like Insertion Sequences in Pseudomonas stutzeri after Conjugative Interaction
TnpR Encoded by an ISPpu12 Isoform Regulates Transposition of Two Different ISL3-Like Insertion Sequences in Pseudomonas stutzeri after Conjugative Interaction
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DOI：
10.1128/jb.01336-09
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发表时间：
2010-03-01
影响因子：
3.2
通讯作者：
Bosch, R.
中科院分区：
文献类型：
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作者：
Christie-Oleza, J. A.;Nogales, B.;Bosch, R.
施氏假单胞菌AN 10具有两个ISL 3样插入序列（IS）。其中一个最近被描述为ISPst 9。在这项研究中，我们表明，第二个IS，位于4.5 kb上游的ISPst 9，是一个异构体的ISPpu 12从恶臭假单胞菌MT-2。虽然两个ISL 3样IS两侧几乎相同（21/24保守残基）的反向重复序列（IR），并具有相似的转座酶（93%的氨基酸同一性），但它们的伴随基因不同。如对ISPst 9所述，ISPpu 12的同种型也通过保守机制转座，形成环状双链DNA（dsDNA）转座中间体，并且通过与接合菌株大肠杆菌S17-1 λ（pir）相互作用（接合相互作用）而不是与非接合大肠杆菌相互作用来诱导。coli DH5 α。事实上，我们证明，ISPst 9转座后，共轭相互作用发生只有当ISPpu 12存在，表明ISPpu 12上调转座的两个IS在这种条件下。我们还表明，这种接合相互作用介导的诱导ISPpu 12是不排他的P.斯氏疟原虫AN 10菌株，但更普遍的现象，至少在假单胞菌。TnpR是一种存在于ISPpu 12中但不存在于ISPst 9中的MerR样转录调控因子，其突变降低了tnpA（ISPpu 12转座酶编码基因）的转录，并减少了接合相互作用后环状dsDNA转座中间体的形成。TnpR突变体的互补恢复了表型。此外，TnpR在无ISPpu 12遗传背景中的存在在接合相互作用后不诱导ISPst 9。因此，我们的研究结果表明，TnpR，接合相互作用后，激活ISPpu 12的tnpA的转录。然后，ISPpu 12的TnpA将结合ISPpu 12和ISPst 9这两个IS的IR，引起它们的转座。
Pseudomonas stutzeri AN10 has two ISL3-like insertion sequences (ISs). One of them has been recently described as ISPst9. In this study we show that the second IS, situated 4.5 kb upstream of ISPst9, is an isoform of ISPpu12 from Pseudomonas putida mt-2. Although both ISL3-like ISs are flanked by nearly identical (21/24 conserved residues) inverted repeats (IRs) and harbor similar transposases (93% amino acid identity), they differ in their accompanying genes. As described for ISPst9, the isoform of ISPpu12 also transposes by a conservative mechanism, forms circular double-stranded DNA (dsDNA) transposition intermediates, and is induced by interaction with the conjugative strain Escherichia coli S17-1 lambda(pir) (conjugative interaction) but not with the nonconjugative E. coli DH5 alpha. In fact, we demonstrate that ISPst9 transposition after conjugative interaction occurs only when ISPpu12 is present, indicating that ISPpu12 is upregulating transposition of both ISs under such conditions. We also demonstrate that this conjugative interaction-mediated induction of ISPpu12 is not exclusive to the P. stutzeri AN10 strain but is a more general phenomenon, at least in Pseudomonas. Mutation of TnpR, a MerR-like transcriptional regulator present in ISPpu12 but not in ISPst9, reduced the transcription of tnpA (ISPpu12 transposase-encoding gene) and decreased formation of circular dsDNA transposition intermediates after conjugative interaction. Complementation of the TnpR mutant restored the phenotype. In addition, the presence of TnpR in an ISPpu12-free genetic background did not induce ISPst9 after conjugative interaction. Thus, our results suggest that TnpR, after conjugative interaction, activates transcription of tnpA of ISPpu12. Then, TnpA of ISPpu12 would bind to IRs of both ISs, ISPpu12 and ISPst9, causing their transposition.