Terceti Mateus S, Rivas Amable J, Alvarez Laura, Noia Manuel, Cava Felipe, Osorio Carlos R
Departamento de Microbioloxía e Parasitoloxía, Instituto de Acuicultura, Universidade de Santiago de CompostelaSantiago de Compostela, Spain.
Department of Molecular Biology and Laboratory for Molecular Infection Medicine Sweden, Umeå Centre for Microbial Research, Umeå UniversityUmeå, Sweden.
Front Microbiol. 2017 Apr 10;8:582. doi: 10.3389/fmicb.2017.00582. eCollection 2017.
The marine pathogenic bacterium subsp. causes septicemia in marine animals and in humans. The pPHDD1 plasmid-encoded hemolysins damselysin (Dly) and phobalysin P (PhlyP), and the chromosome-encoded hemolysin phobalysin C (PhlyC) constitute its main virulence factors. However, the mechanisms by which expression of these three hemolysins is regulated remain unknown. Here we report the isolation of a mini-Tn transposon mutant which showed a strong impairment in its hemolytic activity. The transposon disrupted a putative sensor histidine kinase gene (), which together with () is predicted to encode a putative two-component regulatory system. This system showed to be homologous to the CarSR/VprAB and RstAB systems. Reconstruction of the mutant by allelic exchange of showed equal impairment in hemolysis, and complementation with a plasmid expressing restored hemolysis to wild-type levels. Remarkably, we demonstrated by promoter expression analyses that the reduced hemolysis in the mutant was accompanied by a strong decrease in transcription activities of the three hemolysin genes (damselysin), (phobalysin P) and (phobalysin C). Thus, RstB, encoded in the small chromosome, regulates plasmid and chromosomal virulence genes. We also found that reduced expression of the three virulence genes correlated with a strong decrease in virulence in a sea bass model, demonstrating that RstB constitutes a master regulator of the three subsp. hemolysins and plays critical roles in the pathogenicity of this bacterium. This study represents the first evidence of a direct role of a RstAB-like system in the regulation of bacterial toxins.
海洋致病细菌亚种会导致海洋动物和人类患败血症。pPHDD1质粒编码的溶血素damselysin（Dly）和phobalysin P（PhlyP），以及染色体编码的溶血素phobalysin C（PhlyC）构成其主要毒力因子。然而，这三种溶血素的表达调控机制仍不清楚。在此，我们报告了一个mini-Tn转座子突变体的分离，该突变体在溶血活性方面表现出严重受损。转座子破坏了一个假定的传感组氨酸激酶基因（），该基因与（）一起预计编码一个假定的双组分调节系统。该系统显示与CarSR/VprAB和RstAB系统同源。通过等位基因交换对突变体进行重建显示溶血功能同样受损，用表达的质粒进行互补可将溶血恢复到野生型水平。值得注意的是，我们通过启动子表达分析证明，突变体中溶血减少伴随着三个溶血素基因（damselysin）、（phobalysin P）和（phobalysin C）转录活性的强烈下降。因此，小染色体中编码的RstB调节质粒和染色体毒力基因。我们还发现，在海鲈模型中，这三个毒力基因表达的降低与毒力的强烈下降相关，表明RstB构成了亚种三种溶血素的主要调节因子，并在该细菌的致病性中起关键作用。这项研究首次证明了RstAB样系统在细菌毒素调节中的直接作用。
