Environmental regulation of gene expression dissected by microfluidics

描述（由申请人提供）：人牙菌斑生物膜是一个物理和化学复杂的环境，其中居住着许多细菌物种，包括变形链球菌，变形链球菌被认为是人龋齿的主要病因。S.变形杆菌有许多导致其毒性的行为，它通过使用从其环境中接收的化学线索来调节和激活这些行为。然而，S.在生物膜内不同位置处的变形菌可能在化学和物理上不同，受到pH、氧浓度和营养物可用性的实质性变化，以及定植物种的不同平衡和允许细菌交流和调节毒力行为的不同浓度的小分子的影响。这个项目旨在确定微环境的多样性如何影响重要的S。变异株的毒力行为称为遗传能力。感受态是指S.变形杆菌与调节生物体的几乎所有其他致龋特征的机制密切相关。感受态基因对环境条件和所接收的化学信号的性质极其敏感。感受态基因在整个细胞群中的表达可以是均匀的，或者可以涉及仅激活细菌的一个子集。因此，一个广泛的科学目标是了解S。变形杆菌处理或解释环境信号以调节感受态和其它毒力行为，生物膜中的微环境如何影响该处理，以及感受态基因如何在整个口腔生物膜中在细胞-细胞水平上被激活。本计画将著重于辨识与了解控制沙门氏菌的基因“开关”。变形链球菌的感受态，探讨pH、氧浓度和碳水化合物的可利用性如何影响肽信号对感受态的调节，并研究和模拟了变形链球菌内肽信号对感受态的调节方式。变形菌生物膜该项目将通过一种新的微流体单细胞方法来实现这些目标。这种方法允许多个，明确定义的环境输入提供给亚群的S。变形细胞，同时测量跨这些亚群的感受态基因激活的概况并定量建模。该项目将产生有关S.口腔生物膜中的变异体感受态是空间分布的，以及什么样的化学条件触发了这种感受态和相关的毒力行为。成功实现这些目标将通过提高对变形链球菌如何引起牙科疾病的理解来推进人类口腔健康的研究。

DESCRIPTION (provided by applicant): The human dental plaque biofilm is a physically and chemically complex environment that is inhabited by a many bacterial species, including Streptococcus mutans, which is regarded as the primary etiological cause of human dental caries. S. mutans has a number of behaviors that give rise to its virulence, and it regulates and activates these behaviors through the use of chemical cues received from its environment. However, the local environment of S. mutans at different locations within a biofilm may be chemically and physically different, subject to substantial variation in pH, oxygen concentration and nutrient availability, as well as different balances of colonizing species and different concentrations of the small molecules that allow the bacteria to communicate and regulate virulence behaviors. This project seeks to determine how this diversity of microenvironments affects the centrally important S. mutans virulence behavior known as genetic competence. Competence is the ability to take up extracellular DNA, and the genetic network that regulates competence in S. mutans is closely intertwined with the mechanisms that regulate virtually every other cariogenic trait of the organism. The competence genes are extremely sensitive to environmental conditions and to the nature of the chemical signals received. Expression of competence genes across a population of cells can be uniform or can involve activation of only a subset of the bacteria. Therefore a broad scientific goal is to understand how S. mutans processes or interprets environmental signals to regulate competence and other virulence behaviors, how microenvironments in the biofilm affect this processing, and how competence genes are activated at the cell-to-cell level throughout an oral biofilm. This project will focus o identifying and understanding the genetic "switches" that control S. mutans competence, exploring how pH, oxygen concentration, and carbohydrate availability affect the regulation of competence by peptide signals, and studying and modeling the ways that competence is regulated by peptide signals inside an S. mutans biofilm. The project will accomplish these goals through a novel microfluidic, single-cell approach. This method allows multiple, well-defined environmental inputs to be supplied to subpopulations of S. mutans cells while the profile of competence gene activation across those subpopulations is measured and modeled quantitatively. The project will yield detailed information about how S. mutans competence is spatially distributed in an oral biofilm and what kinds of chemical conditions trigger this and related virulence behaviors. Success in achieving these objectives will advance research in human oral health by improving the understanding of how Streptococcus mutans causes dental disease.