Department of Molecular Biology, University of Gdańsk, Kładki 24, 80-822 Gdańsk, Poland.
Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Kładki 24, 80-822 Gdańsk, Poland.
Microbiology (Reading). 2011 Jan;157(Pt 1):220-233. doi: 10.1099/mic.0.042820-0. Epub 2010 Sep 9.
The pathogenicity of Shiga toxin-producing Escherichia coli (STEC) depends on the expression of stx genes that are located on lambdoid prophages. Effective toxin production occurs only after prophage induction, and one may presume that replication of the phage genome is important for an increase in the dosage of stx genes, positively influencing their expression. We investigated the replication of plasmids derived from Shiga toxin (Stx)-converting bacteriophages in starved E. coli cells, as starvation conditions may be common in the intestine of infected humans. We found that, unlike plasmids derived from bacteriophage λ, the Shiga toxin phage-derived replicons did not replicate in amino acid-starved relA(+) and relA(-) cells (showing the stringent and relaxed responses to starvation, respectively). The presence of the stable fraction of the replication initiator O protein was detected in all tested replicons. However, while ppGpp, the stringent response effector, inhibited the activities of the λ P(R) promoter and its homologues from Shiga toxin-converting bacteriophages, these promoters, except for λ P(R), were only weakly stimulated by the DksA protein. We suggest that this less efficient (relative to λ) positive regulation of transcription responsible for transcriptional activation of the origin contributes to the inhibition of DNA replication initiation of Shiga toxin-converting bacteriophages in starved host cells, even in the absence of ppGpp (as in starved relA(-) hosts). Possible clinical implications of these results are discussed.
产志贺毒素大肠杆菌（STEC）的致病性取决于位于 lambdoid 噬菌体上的 stx 基因的表达。只有在噬菌体诱导后才能有效产生毒素，并且人们可以假定噬菌体基因组的复制对于 stx 基因剂量的增加很重要，从而对其表达产生积极影响。我们研究了在饥饿的大肠杆菌细胞中源自志贺毒素（Stx）转换噬菌体的质粒的复制，因为感染人类的肠道中可能经常出现饥饿条件。我们发现，与源自噬菌体 λ 的质粒不同，源自志贺毒素噬菌体的复制子在氨基酸饥饿的 relA(+)和 relA(-)细胞中（分别表现出严格和松弛的饥饿反应）不复制。在所有测试的复制子中都检测到复制起始蛋白 O 的稳定部分。然而，虽然 ppGpp（严格反应效应物）抑制了 λ P(R)启动子及其源自志贺毒素转换噬菌体的同源物的活性，但这些启动子除了 λ P(R)之外，仅被 DksA 蛋白弱刺激。我们认为，这种转录的正调控效率较低（相对于 λ），负责转录起始的起始有助于抑制噬菌体在饥饿宿主细胞中的 DNA 复制起始，即使在没有 ppGpp 的情况下（如在饥饿的 relA(-)宿主中）。讨论了这些结果的可能临床意义。
