Mohaisen Mohammed Radhi, McCarthy Alan John, Adriaenssens Evelien M, Allison Heather Elizabeth
Department of Functional and Comparative Genomics, Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom.
College of Dentistry, University of Anbar, Ramadi, Iraq.
Front Microbiol. 2020 Oct 9;11:578056. doi: 10.3389/fmicb.2020.578056. eCollection 2020.
Stx bacteriophages are members of the lambdoid group of phages and are responsible for Shiga toxin (Stx) production and the dissemination of Shiga toxin genes () across shigatoxigenic (STEC). These toxigenic bacteriophage hosts can cause life-threatening illnesses, and Stx is the virulence determinant responsible for the severe nature of infection with enterohemorrhagic , a subset of pathogenic STEC. Stx phages are temperate, and in the present study, the identification of what is actually required for Stx phage Φ24 and bacterial DNA recombination was tested using both and recombination assays. It is well established that phage λ, which underpins most of what we understand about lambdoid phage biology, requires its own encoded phage attachment site () of 250 bp, a host-encoded attachment site () of 21 bp, and a host-encoded DNA binding protein known as integration host factor (IHF). The assays applied in this study enabled the manipulation of the phage attachment site () and the bacterial attachment site () sequences and the inclusion or exclusion of a host-encoded accessory element known as integration host factor. We were able to demonstrate that the minimal sequence required by Φ24 phage is between 350 and 427 bp. Unlike phage λ, the minimal necessary flanking sequences for the site do not appear to be equal in size, with a total length between 62 and 93 bp. Furthermore, we identified that the Φ24 integrase does not require IHF to drive the integration and the recombination process. Understanding how this unusual Stx phage integrase works may enable exploitation of its promiscuous nature in the context of genetic engineering.
志贺毒素噬菌体是λ样噬菌体群的成员，负责志贺毒素（Stx）的产生以及志贺毒素基因在产志贺毒素大肠杆菌（STEC）中的传播。这些产毒噬菌体宿主可导致危及生命的疾病，而Stx是导致肠出血性大肠杆菌（致病性STEC的一个子集）感染严重性的毒力决定因素。Stx噬菌体是温和噬菌体，在本研究中，使用λ和Rac重组试验测试了Stx噬菌体Φ24和细菌DNA重组实际所需的条件。众所周知，支撑我们对λ样噬菌体生物学大部分理解的噬菌体λ，需要其自身编码的250 bp噬菌体附着位点（attP）、21 bp的宿主编码附着位点（attB）以及一种称为整合宿主因子（IHF）的宿主编码DNA结合蛋白。本研究中应用的试验能够操纵噬菌体附着位点（attP）和细菌附着位点（attB）序列，并纳入或排除一种称为整合宿主因子的宿主编码辅助元件。我们能够证明，Φ24噬菌体所需的最小attP序列在350至427 bp之间。与噬菌体λ不同，attB位点所需的最小侧翼序列大小似乎不相等，总长度在62至93 bp之间。此外，我们发现Φ24整合酶不需要IHF来驱动整合和重组过程。了解这种不同寻常的Stx噬菌体整合酶的工作方式，可能有助于在基因工程背景下利用其混杂特性。
