Intra- and inter species cell-cell communication in Vibrio cholerae

描述(申请人提供)：人类病原体霍乱弧菌使用群体感应，涉及生产、释放和随后检测称为自动诱导物的化学信号分子，以调节毒力基因的表达，以应对细胞种群密度和物种复杂性的变化。目前已知两种霍乱弧菌自动诱导检测系统，一种用于种内，另一种用于种间交流。这项应用的长期目标是了解这种细菌中潜在的种内和种间群体感应信号的分子机制。编码在每个细胞外自身诱导子中的唯一信息在内部被翻译成磷酸化反应调节器Luxo的离散水平，该调节器决定霍乱弧菌群体传感系统的最终输出。由于天冬氨酸磷酸酯的不稳定性，在体内的磷酸化细菌反应调节剂的水平从未被直接测定过。为了了解霍乱弧菌群体感应电路中的信号-响应输入-输出关系，我的第一个目标是建立定量测量霍乱弧菌细胞中磷-Luxo水平的方法学。将两个不同的自动诱导子中包含的信息整合到一个单一的调控网络中，可以使霍乱弧菌群体感应系统发挥符合检测器的作用。或者，这个回路可以通过两个自动诱导子的任何或所有组合来促进不同的基因调控。我的第二个目标是通过研究每个自体诱导剂对特定靶基因表达的影响来区分这两种可能性。将对所选目标基因的时间表达动态进行测量，以确定自动诱导子是否共同发挥作用，或者是否存在响应不同自动诱导剂组合的有序基因表达层次。这些研究的结果将充分确定每个调节元件在群体感应信号通路中的作用。这项应用的第三个目的是识别存在于霍乱弧菌和其他细菌物种之间的新的化学信号通信途径。物种间的交流可能使霍乱弧菌能够适应、竞争并在复杂的环境中生存，例如受污染的水和存在许多其他细菌物种的人体肠道。将确定和描述这种物种间交流途径所涉及的组成部分。总之，这些研究将为霍乱弧菌种内和种间通信网络的分子结构提供洞察力。实际上，这项工作的结果可能会导致反群体感应疗法的开发，作为传统抗生素的替代品。

DESCRIPTION (provided by applicant): The human pathogen Vibrio cholerae uses quorum sensing which involves the production, release, and subsequent detection of chemical signaling molecules called autoinducers, to regulate virulence gene expression in response to changes in cell-population density and species complexity. Two V. cholerae autoinducer-detection systems, one for intra-species and one for inter-species communication, are currently known. The long term goal of this application is to understand the molecular mechanisms underlying intra-species and inter-species quorum-sensing signaling in this bacterium. The unique information encoded in each extracellular autoinducer is translated internally into discrete levels of the phosphorylated response regulator LuxO, which determines the final output of the V. cholerae quorum-sensing system. Due to the instability of aspartyl phosphates, in vivo levels of phosphorylated bacterial response regulators have never been directly determined. To understand the signal-response input-output relationship in the V. cholerae quorum-sensing circuit, my first aim is to develop methodology for the quantitative measurement of phospho-LuxO levels in V. cholerae cells. Integration of the information contained in the two different autoinducers into a single regulatory network could allow the V. cholerae quorum-sensing system to function as a coincidence detector. Alternatively, this circuit may facilitate distinct gene regulation by any or all combinations of the two autoinducers. My second aim is focused on distinguishing between these two possibilities by investigating the effect of each autoinducer on specific target gene expression. Measurement of the temporal expression dynamics of selected target genes will be performed to establish whether the autoinducers function together or if there is an ordered gene expression hierarchy in response to different autoinducer combinations. Results from these studies will fully define the role of each regulatory element in the quorum-sensing signaling circuit. The third aim of this application is to identify novel chemical signal communication pathways that exist between V. cholerae and other bacterial species. Inter-species communication likely enables V. cholerae to adapt, compete, and survive in complex environments such as contaminated water and the human intestine where many other bacterial species are present. The components involved in such inter-species communication pathways will be identified and characterized. Together, these studies will provide insight into the molecular architecture of V. cholerae intra- and inter-species communication networks. Practically, results from this work could lead to the development of anti-quorum-sensing therapies for use as alternatives to traditional antibiotics.