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首页> 外文期刊>Journal of bacteriology >Role of hha and ler in Transcriptional Regulation of the esp Operon of Enterohemorrhagic Escherichia coli O157:H7
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Role of hha and ler in Transcriptional Regulation of the esp Operon of Enterohemorrhagic Escherichia coli O157:H7

机译：hha和ler在肠出血性大肠杆菌O157：H7的esp操纵子转录调控中的作用

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The locus of enterocyte effacement (LEE), which includes five major operons (LEE1 through LEE4 and tir), enables enterohemorrhagic Escherichia coli (EHEC) O157:H7 to produce attaching and effacing lesions on host cells. Expression of LEE2, LEE3, and tir is positively regulated by ler, a gene located in LEE1. Transcriptional regulation of the esp operon (LEE4), however, is not well defined. Transposon mutagenesis was used to identify transcriptional regulators of the esp operon by screening for mutants with increased β-galactosidase activity in an EHEC O157:H7 strain harboring an esp::lac transcriptional fusion. All mutants with significant increases in β-galactosidase activity had transposon insertions in hha (hha::Tn). Specific complementation of the hha::Tn mutation with a plasmid-encoded copy of hha reduced β-galactosidase activity to the level expressed in the parental esp::lac strain. Purified Hha, however, bound poorly to the esp promoter, suggesting that Hha might repress the transcription of a positive regulator of esp. Transposon mutagenesis of a Δhha esp::lac strain expressing elevated levels of β-galactosidase resulted in ler mutants with reduced β-galactosidase activity. Purified Hha bound to the ler promoter with a higher affinity, and complementation of a Δhha mutation in a Δhha ler::lac strain repressed β-galactosidase activity to the level expressed in a ler::lac strain. A positive regulatory role of ler in esp expression was demonstrated by specific binding of Ler to the esp promoter, reduced expression of β-galactosidase in Δler esp::lac strains with and without hha, and severalfold-increased transcription of ler and espA in strains lacking hha. These results indicate that hha-mediated repression of ler causes reduced expression of the esp operon.
机译：肠细胞外溢（LEE）的位点包括五个主要操纵子（ LEE1 至 LEE4 和 tir ），可使肠出血性大肠杆菌（EHEC）O157：H7在宿主细胞上产生附着和脱落的损伤。 LEE2 ， LEE3 和 tir 的表达受到 ler 的正调控，该基因位于 LEE1 。然而，对 esp 操纵子（ LEE4 ）的转录调控尚不明确。转座子诱变通过在带有 esp ::: lac 转录融合。 β-半乳糖苷酶活性显着增加的所有突变体在 hha （ hha :: Tn）中都有转座子插入。 hha :: Tn突变与 hha 的质粒编码拷贝的特异性互补将β-半乳糖苷酶活性降低到亲本 esp :: lac 菌株。但是，纯化的Hha与 esp 启动子的结合较弱，表明Hha可能抑制了 esp 阳性调节子的转录。 β-半乳糖苷酶水平升高的Δ hha esp :: lac 菌株的转座子诱变导致 ler 突变体的β-半乳糖苷酶活性降低。纯化的Hha以更高的亲和力与 ler 启动子结合，并互补Δ hha ler :: 中的Δ hha 突变lac 株将β-半乳糖苷酶活性抑制到 ler :: lac 株中表达的水平。 Ler与 esp 启动子的特异性结合，减少了Δ中β-半乳糖苷酶的表达，证明了 ler 在 esp 表达中的正调节作用。带有和不带有 hha 的 ler esp :: lac 菌株， ler 和的转录倍数增加espA 在缺乏 hha 的菌株中。这些结果表明 hha 介导的 ler 阻遏导致 esp 操纵子的表达减少。

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