韩雨, 黄元铭, 程倩, 李杰, 刁保卫, 梁未丽. 河弧菌VfqI-VfqR密度感应系统对VflT6SS2附属基因簇中vgrG操纵子的调控研究[J]. 疾病监测, 2022, 37(5): 585-590. DOI: 10.3784/jbjc.202203180100
引用本文: 韩雨, 黄元铭, 程倩, 李杰, 刁保卫, 梁未丽. 河弧菌VfqI-VfqR密度感应系统对VflT6SS2附属基因簇中vgrG操纵子的调控研究[J]. 疾病监测, 2022, 37(5): 585-590. DOI: 10.3784/jbjc.202203180100
Han Yu, Huang Yuanming, Cheng Qian, Li Jie, Diao Baowei, Liang Weili. Regulation of VfqI-VfqR Quorum sensing circuit on vgrG operon in VflT6SS orphan cluster of Vibrio fluvialis[J]. Disease Surveillance, 2022, 37(5): 585-590. DOI: 10.3784/jbjc.202203180100
Citation: Han Yu, Huang Yuanming, Cheng Qian, Li Jie, Diao Baowei, Liang Weili. Regulation of VfqI-VfqR Quorum sensing circuit on vgrG operon in VflT6SS orphan cluster of Vibrio fluvialis[J]. Disease Surveillance, 2022, 37(5): 585-590. DOI: 10.3784/jbjc.202203180100

河弧菌VfqI-VfqR密度感应系统对VflT6SS2附属基因簇中vgrG操纵子的调控研究

Regulation of VfqI-VfqR Quorum sensing circuit on vgrG operon in VflT6SS orphan cluster of Vibrio fluvialis

  • 摘要:
      目的  探究河弧菌VfqI-VfqR密度感应系统对VflT6SS2附属基因簇hcp-vgrGvgrG操纵子的调控。
      方法  使用荧光定量反转录聚合酶链式反应(qRT-PCR)检测野生株和VfqI-VfqR系统缺失株中vgrG的mRNA水平。 利用启动子-lux报告质粒系统检测冷光值,确定vgrG启动子在河弧菌野生株、缺失株、回补株中的活性差异。 在大肠埃希菌中,导入VfqR过表达质粒,检测vgrG启动子-lux报告质粒冷光值,确定VfqR可否激活vgrG启动子。
      结果  ∆vfqI、∆vfqR和∆vfqI-vfqR缺失株中vgrG mRNA水平和启动子活性均显著低于野生株;在∆vfqR中回补VfqR表达质粒pSRvfqR和在∆vfqI中分别添加3-oxo-C10-HSL、C10-HSL和3-oxo-C12-HSL AHLs分子均能恢复vgrG启动子活性,并且3种AHLs分子效果大体一致。 在大肠埃希菌中过表达VfqR并添加上述3种外源AHLs,对vgrG启动子活性没有影响。
      结论  VfqI-VfqR密度感应系统在启动子水平正性调控vgrG操纵子,并且该调控作用是间接的。

     

    Abstract:
      Objective  To explore the regulation of the vgrG operon in the VflT6SS2 orphan clusters by the Quorum sensing system VfqI-VfqR of Vibrio fluvialis.
      Methods  Quantitative reverse transcription PCR was used to detect the relative expression of vgrG mRNA level in wild V. fluvialis strain and VfqI-VfqR system deletion mutant strains. Luminescence of promoter-lux fusion was measured to determine the vgrG promoter activity in wild V. fluvialis strain, derivative mutants and corresponding complemented strains. VfqR-overexpressed plasmid was introduced into Escherichia coli, the luminescence of vgrG promoter-lux fusion was compared to determine the direct effect of VfqR on vgrG operon activation.
      Results  The vgrG mRNA and its promoter activity significantly decreased in the VfqI-VfqR deletion mutants compared with wild V. fluvialis strain. Furthermore, the vgrG promoter activity of ∆vfqR was restored either with trans-complemented plasmid or supplemented with exogenous 3-oxo-C10-HSL, C10-HSL or 3-oxo-C12-HSL AHL molecules. The effects of the three AHLs were similar. However, overexpression of VfqR combined with addition of AHLs had no effect on the promoter activity of vgrG in E. coli.
      Conclusion  The VfqI-VfqR system is one of positive regulators of vgrG operator indirectly tuning the activation of its promoter.

     

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