Roles for five small RNAs required for quorum sensing in Vibrio harveyi

描述(由申请者提供)：我的首要目标是了解支撑集体行为的分子机制。为了探索这一点，我将研究群体感应，这是模式细菌哈维氏弧菌中细胞间交流的过程：利用群体感应，细菌产生、检测和响应细胞外信号分子(自动诱导物)，以在种群范围内协调基因表达。在V.harveyi中，五个调节的小RNA(称为QrR sRNAs)在低细胞密度下产生，并作用于破坏编码主要群体感应调节因子LuxR的LuxR mRNA的稳定。我的目标是了解具有五个QrR sRNA而不是一个sRNA的哪些功能提供了哈维氏弧菌的群体感应电路，以及更广泛地说，sRNA而不是调节蛋白为信号网络提供了哪些功能。具体地说，我将解决以下几点。1)确定分别控制每个QRR sRNA基因的转录因子。5个c/rrsRNA基因启动子存在差异，每个gfrrsRNA基因表现出不同的基因表达模式，提示不同的转录因子调控着每个qrrsRNA基因的表达。我建议鉴定这些转录因子，并研究它们在哈维氏弧菌群体感应反应中的作用。2)确定每个Qrr sRNA所独有的信使核糖核酸靶点。我将确定由每个哈维氏弧菌QrrsRNA唯一调控的基因，并确定确定的靶标的群体感应功能。3)确定靶基因的不同QrR SRNA亲和力如何建立群体感应基因表达的有序程序。我认为QrR sRNA与mRNA靶标结合的亲和力不同，这会影响各种群体感应反应输出的时间。我将比较QrR sRNA与已知的哈维氏弧菌mRNA靶标的结合强度，定义QrR sRNA结合的序列要求，以及改变这些结合特性如何影响群体感应基因表达的时间。我们的群体感应研究的一个重要的实际方面是，它们正在导致反群体感应疗法的发展，以用作传统抗生素的替代品。同样，对有益细菌群体感应的分子理解，例如那些生产重要医疗、农业或工业产品的细菌，可以促进群体感应战略的开发和实施，从而改善具有商业意义的天然产品的工业规模生产。

DESCRIPTION (provided by applicant): My overarching goal is to understand the molecular mechanisms underpinning collective behaviors. To explore this, I will study quorum sensing, the process of cell-cell communication in the model bacterium Vibrio harveyi: Using quorum sensing, bacteria produce, detect, and respond to extracellular signal molecules (autoinducers) to coordinate gene expression on a population-wide scale. In V. harveyi, five regulatory small RNAs (called Qrr sRNAs) are produced at low cell densities and act to destabilize the luxR mRNA encoding the master quorum-sensing regulator, LuxR. My goal is to understand what features having five Qrr sRNAs rather than one sRNA provides the V. harveyi quorum-sensing circuit, and, more generally, what features sRNAs rather than regulatory proteins provide to signaling networks. Specifically, I will address the following points. 1) Define the transcription factors that individually control each qrr sRNA gene. The five c/rr sRNA gene promoters are divergent, and each gfrrsRNA gene shows a distinct pattern of gene expression, suggesting that different transcription factors regulate expression of each Qrr sRNA gene. I propose to identify these transcription factors and characterize their roles in the V. harveyi quorum-sensing response. 2) Identify mRNA targets that are exclusive to each Qrr sRNA. I will identify genes regulated exclusively by each of the V. harveyi Qrr sRNAs and determine the quorum-sensing functions of the identified targets. 3) Determine how different Qrr sRNA affinities for target mRNAs establish an ordered program of quorum-sensing gene expression. I propose that the Qrr sRNAs bind with different affinities to mRNA targets and this affects the timing of the various quorum-sensing response outputs. I will compare binding strengths of the Qrr sRNAs with known V. harveyi mRNA targets, define the sequence requirements for Qrr sRNA binding, and how altering those binding properties affects the timing of quorum- sensing gene expression. An important practical aspect of our quorum-sensing investigations is that they are leading to the development of anti-quorum-sensing therapies for use as alternatives to traditional antibiotics. Similarly, a molecular understanding of quorum sensing in beneficial bacteria, such as those that produce medical, agricultural, or industrial products of importance, could facilitate the development and implementation of pro- quorum-sensing strategies that improve industrial-scale production of commercially relevant natural products.