Pilla Giulia, McVicker Gareth, Tang Christoph M
Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom.
PLoS Genet. 2017 Sep 25;13(9):e1007014. doi: 10.1371/journal.pgen.1007014. eCollection 2017 Sep.
Acquisition of a single copy, large virulence plasmid, pINV, led to the emergence of Shigella spp. from Escherichia coli. The plasmid encodes a Type III secretion system (T3SS) on a 30 kb pathogenicity island (PAI), and is maintained in a bacterial population through a series of toxin:antitoxin (TA) systems which mediate post-segregational killing (PSK). The T3SS imposes a significant cost on the bacterium, and strains which have lost the plasmid and/or genes encoding the T3SS grow faster than wild-type strains in the laboratory, and fail to bind the indicator dye Congo Red (CR). Our aim was to define the molecular events in Shigella flexneri that cause loss of Type III secretion (T3S), and to examine whether TA systems exert positional effects on pINV. During growth at 37°C, we found that deletions of regions of the plasmid including the PAI lead to the emergence of CR-negative colonies; deletions occur through intra-molecular recombination events between insertion sequences (ISs) flanking the PAI. Furthermore, by repositioning MvpAT (which belongs to the VapBC family of TA systems) near the PAI, we demonstrate that the location of this TA system alters the rearrangements that lead to loss of T3S, indicating that MvpAT acts both globally (by reducing loss of pINV through PSK) as well as locally (by preventing loss of adjacent sequences). During growth at environmental temperatures, we show for the first time that pINV spontaneously integrates into different sites in the chromosome, and this is mediated by inter-molecular events involving IS1294. Integration leads to reduced PAI gene expression and impaired secretion through the T3SS, while excision of pINV from the chromosome restores T3SS function. Therefore, pINV integration provides a reversible mechanism for Shigella to circumvent the metabolic burden imposed by pINV. Intra- and inter-molecular events between ISs, which are abundant in Shigella spp., mediate plasticity of S. flexneri pINV.
获得单个拷贝的大型毒力质粒pINV导致了志贺氏菌属从大肠杆菌中出现。该质粒在一个30 kb的致病岛（PAI）上编码III型分泌系统（T3SS），并通过一系列毒素：抗毒素（TA）系统在细菌群体中得以维持，这些系统介导分离后杀伤（PSK）。T3SS给细菌带来了巨大代价，在实验室中，丢失质粒和/或编码T3SS的基因的菌株比野生型菌株生长得更快，并且无法结合指示染料刚果红（CR）。我们的目标是确定弗氏志贺氏菌中导致III型分泌（T3S）丧失的分子事件，并研究TA系统是否对pINV产生位置效应。在37°C生长期间，我们发现质粒包括PAI区域的缺失会导致CR阴性菌落的出现；缺失是通过PAI侧翼插入序列（ISs）之间的分子内重组事件发生的。此外，通过将MvpAT（属于TA系统的VapBC家族）重新定位到PAI附近，我们证明该TA系统的位置改变了导致T3S丧失的重排，表明MvpAT既在全局发挥作用（通过减少pINV因PSK而丢失），也在局部发挥作用（通过防止相邻序列丢失）。在环境温度下生长期间，我们首次表明pINV会自发整合到染色体的不同位点，这是由涉及IS1294的分子间事件介导的。整合导致PAI基因表达降低以及通过T3SS的分泌受损，而从染色体上切除pINV可恢复T3SS功能。因此，pINV整合为志贺氏菌规避pINV带来的代谢负担提供了一种可逆机制。志贺氏菌属中丰富的ISs之间的分子内和分子间事件介导了弗氏志贺氏菌pINV的可塑性。
