Global Health Institute, School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne-EPFL, Lausanne, Switzerland.
PLoS Genet. 2012;8(6):e1002778. doi: 10.1371/journal.pgen.1002778. Epub 2012 Jun 21.
The human pathogen Vibrio cholerae is an aquatic bacterium frequently encountered in rivers, lakes, estuaries, and coastal regions. Within these environmental reservoirs, the bacterium is often found associated with zooplankton and more specifically with their chitinous exoskeleton. Upon growth on such chitinous surfaces, V. cholerae initiates a developmental program termed "natural competence for genetic transformation." Natural competence for transformation is a mode of horizontal gene transfer in bacteria and contributes to the maintenance and evolution of bacterial genomes. In this study, we investigated competence gene expression within this organism at the single cell level. We provide evidence that under homogeneous inducing conditions the majority of the cells express competence genes. A more heterogeneous expression pattern was observable on chitin surfaces. We hypothesize that this was the case due to the heterogeneity around the chitin surface, which might vary extensively with respect to chitin degradation products and autoinducers; these molecules contribute to competence induction based on carbon catabolite repression and quorum-sensing pathways, respectively. Therefore, we investigated the contribution of these two signaling pathways to natural competence in detail using natural transformation assays, transcriptional reporter fusions, quantitative RT-PCR, and immunological detection of protein levels using Western blot analysis. The results illustrate that all tested competence genes are dependent on the transformation regulator TfoX. Furthermore, intracellular cAMP levels play a major role in natural transformation. Finally, we demonstrate that only a minority of genes involved in natural transformation are regulated in a quorum-sensing-dependent manner and that these genes determine the fate of the surrounding DNA. We conclude with a model of the regulatory circuit of chitin-induced natural competence in V. cholerae.
人类病原体霍乱弧菌是一种水生细菌，经常在河流、湖泊、河口和沿海地区发现。在这些环境储库中，该细菌通常与浮游动物有关，更具体地说，与它们的几丁质外骨骼有关。在这些几丁质表面生长时，霍乱弧菌会启动一个称为“天然遗传转化能力”的发育程序。天然转化能力是细菌中一种水平基因转移的模式，有助于维持和进化细菌基因组。在这项研究中，我们在单细胞水平上研究了该生物体中的竞争基因表达。我们提供的证据表明，在同质诱导条件下，大多数细胞表达竞争基因。在几丁质表面上可以观察到更异质的表达模式。我们假设，由于几丁质表面周围的异质性，可能会在几丁质降解产物和自诱导物方面发生广泛变化，这些分子分别根据碳分解代谢物抑制和群体感应途径有助于竞争诱导。因此，我们使用天然转化测定、转录报告融合、定量 RT-PCR 和 Western blot 分析免疫检测蛋白水平详细研究了这两种信号通路对天然竞争的贡献。结果表明，所有测试的竞争基因都依赖于转化调节剂 TfoX。此外，细胞内 cAMP 水平在天然转化中起着重要作用。最后，我们证明只有少数参与天然转化的基因以群体感应依赖性方式受到调节，这些基因决定了周围 DNA 的命运。我们以霍乱弧菌中几丁质诱导的天然竞争的调控回路模型结束。
