Carraro Nicolas, Durand Romain, Rivard Nicolas, Anquetil Charley, Barrette Catherine, Humbert Malika, Burrus Vincent
Département de biologie, Faculté des sciences, Université de Sherbrooke, Sherbrooke, Québec, Canada.
PLoS Genet. 2017 Mar 29;13(3):e1006705. doi: 10.1371/journal.pgen.1006705. eCollection 2017 Mar.
IncC conjugative plasmids and Salmonella genomic island 1 (SGI1) and relatives are frequently associated with multidrug resistance of clinical isolates of pathogenic Enterobacteriaceae. SGI1 is specifically mobilized in trans by IncA and IncC plasmids (commonly referred to as A/C plasmids) following its excision from the chromosome, an event triggered by the transcriptional activator complex AcaCD encoded by these helper plasmids. Although SGI1 is not self-transmissible, it carries three genes, traNS, traHS and traGS, coding for distant homologs of the predicted mating pore subunits TraNC, TraHC and TraGC, respectively, encoded by A/C plasmids. Here we investigated the regulation of traNS and traHGS and the role of these three genes in the transmissibility of SGI1. Transcriptional fusion of the promoter sequences of traNS and traHGS to the reporter gene lacZ confirmed that expression of these genes is inducible by AcaCD. Mating experiments using combinations of deletion mutants of SGI1 and the helper IncC plasmid pVCR94 revealed complex interactions between these two mobile genetic elements. Whereas traNC and traHGC are essential for IncC plasmid transfer, SGI1 could rescue null mutants of each individual gene revealing that TraNS, TraHS and TraGS are functional proteins. Complementation assays of individual traC and traS mutants showed that not only do TraNS/HS/GS replace TraNC/HC/GC in the mating pore encoded by IncC plasmids but also that traGS and traHS are both required for SGI1 optimal transfer. In fact, remodeling of the IncC-encoded mating pore by SGI1 was found to be essential to enhance transfer rate of SGI1 over the helper plasmid. Furthermore, traGS was found to be crucial to allow DNA transfer between cells bearing IncC helper plasmids, thereby suggesting that by remodeling the mating pore SGI1 disables an IncC-encoded entry exclusion mechanism. Hence traS genes facilitate the invasion by SGI1 of cell populations bearing IncC plasmids.
IncC 接合质粒以及沙门氏菌基因组岛1（SGI1）及其相关元件常与致病性肠杆菌科临床分离株的多药耐药性相关。SGI1从染色体上切除后，会被IncA和IncC质粒（通常称为A/C质粒）特异性地进行反式动员，这一事件由这些辅助质粒编码的转录激活复合物AcaCD触发。虽然SGI1本身不能自我传递，但它携带三个基因，即traNS、traHS和traGS，分别编码与A/C质粒编码的预测性交配孔亚基TraNC、TraHC和TraGC远缘同源的蛋白。在此，我们研究了traNS和traHGS的调控以及这三个基因在SGI1可传递性中的作用。将traNS和traHGS的启动子序列与报告基因lacZ进行转录融合，证实这些基因的表达可被AcaCD诱导。使用SGI1缺失突变体与辅助IncC质粒pVCR94的组合进行接合实验，揭示了这两个可移动遗传元件之间复杂的相互作用。虽然TraNC和TraHGC对IncC质粒转移至关重要，但SGI1能够拯救每个单个基因的缺失突变体，这表明TraNS、TraHS和TraGS是功能性蛋白。对单个traC和traS突变体的互补分析表明，TraNS/HS/GS不仅能在IncC质粒编码的交配孔中替代TraNC/HC/GC，而且traGS和traHS都是SGI1最佳转移所必需的。事实上，发现SGI1对IncC编码的交配孔进行重塑对于提高SGI1相对于辅助质粒的转移率至关重要。此外，发现traGS对于携带IncC辅助质粒的细胞之间的DNA转移至关重要，从而表明通过重塑交配孔，SGI1使IncC编码的进入排斥机制失活。因此，traS基因促进了SGI1对携带IncC质粒的细胞群体的侵入。
