Provenzano D, Klose K E
Department of Microbiology, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, TX 78229-3900, USA.
Proc Natl Acad Sci U S A. 2000 Aug 29;97(18):10220-4. doi: 10.1073/pnas.170219997.
The transmembrane transcriptional activators ToxR and TcpP modulate expression of Vibrio cholerae virulence factors by exerting control over toxT, which encodes the cytoplasmic transcriptional activator of the ctx, tcp, and acf virulence genes. However, ToxR, independently of TcpP and ToxT, activates and represses transcription of the genes encoding two outer-membrane porins, OmpU and OmpT. To determine the role of ToxR-dependent porin regulation in V. cholerae pathogenesis, the ToxR-activated ompU promoter was used to drive ompT transcription in a strain lacking OmpU. Likewise, the ToxR-repressed ompT promoter was used to drive ompU transcription in a strain lacking both ToxR and OmpT. This strategy allowed the generation of a toxR(+) strain that expresses OmpT in place of OmpU, and a toxR(-) strain that expresses OmpU in place of OmpT. Growth rates in the presence of bile salts and other anionic detergents were retarded for the toxR(+) V. cholerae expressing OmpT in place of OmpU, but increased in toxR(-) V. cholerae expressing OmpU in place of OmpT. Additionally, the toxR(+) V. cholerae expressing OmpT in place of OmpU expressed less cholera toxin and toxin-coregulated pilus, and this effect was shown to be caused by reduced toxT transcription in this strain. Finally, the toxR(+) V. cholerae expressing OmpT in place of OmpU was approximately 100-fold reduced in its ability to colonize the infant-mouse intestine. Our results indicate that ToxR-dependent modulation of the outer membrane porins OmpU and OmpT is critical for V. cholerae bile resistance, virulence factor expression, and intestinal colonization.
跨膜转录激活因子ToxR和TcpP通过对toxT进行调控，来调节霍乱弧菌毒力因子的表达，toxT编码ctx、tcp和acf毒力基因的细胞质转录激活因子。然而，ToxR独立于TcpP和ToxT，激活并抑制编码两种外膜孔蛋白OmpU和OmpT的基因的转录。为了确定ToxR依赖性孔蛋白调控在霍乱弧菌致病机制中的作用，在缺乏OmpU的菌株中，使用ToxR激活的ompU启动子来驱动ompT转录。同样，在同时缺乏ToxR和OmpT的菌株中，使用ToxR抑制的ompT启动子来驱动ompU转录。该策略使得能够产生一个用OmpT替代OmpU表达的toxR(+)菌株，以及一个用OmpU替代OmpT表达的toxR(-)菌株。在胆盐和其他阴离子去污剂存在的情况下，用OmpT替代OmpU表达的toxR(+)霍乱弧菌的生长速率受到抑制，但用OmpU替代OmpT表达的toxR(-)霍乱弧菌的生长速率增加。此外，用OmpT替代OmpU表达的toxR(+)霍乱弧菌表达的霍乱毒素和毒素共调节菌毛较少，并且这种效应被证明是由该菌株中toxT转录减少所致。最后，用OmpT替代OmpU表达的toxR(+)霍乱弧菌在定殖于幼鼠肠道的能力上降低了约100倍。我们的结果表明，ToxR依赖性对外膜孔蛋白OmpU和OmpT的调控对于霍乱弧菌的胆汁抗性、毒力因子表达和肠道定殖至关重要。
