3 quorum sensing signals add in microarray comparison in Yersinia pestis at 30°C. Yersinia pestis

The quorum-sensing system has been linked to diverse phenotypes and regulatory changes in pathogenic bacteria. In strain CO92, the AI-2 signal is produced in a luxS-dependent manner, reaching maximal levels of 2.5 μM in late logarithmic growth, and both wild type and pigmentation mutant strains made equivalent levels of AI-2. Yersinia pestis CO92 possesses a chromosomal lsr locus encoding factors involved in the binding and import of AI-2, and confirming this assignment, an lsr deletion increased extracellular pools of AI-2. To assess the functional role of 3 quorum sensing singlas in Y. pestis, microarray study was conducted comparing the 3 quorum sensing signals added in to control at 30°C to mimic the flea gut. Here, the ΔPgm (pigmentation-negative) R88 strain was used. The three signals are one AI-2 and two AHLs (N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone).The control consisted of cells grown and treated under the same conditions without added signals. Overall design: Six independent RNA samples from 3 signals add in cultures were paired with six independent RNA samples from control cultures for hybridization to six two-color microarrays. A dye-swap design was used to remove the Cy5 and Cy3 dye bias.

群体感应系统（quorum-sensing system）已被证实与病原菌的多种表型及调控变化密切相关。在菌株CO92中，AI-2信号以luxS依赖的方式合成，在对数生长后期达到2.5 μM的最高浓度，野生型菌株与色素突变株的AI-2产量相当。鼠疫耶尔森菌（Yersinia pestis）CO92的染色体上携带lsr基因座，其编码参与AI-2结合与摄取的功能因子；lsr基因缺失会导致胞外AI-2池水平升高，这一结果验证了上述功能注释。

为探究鼠疫耶尔森菌中3种群体感应信号的功能作用，研究人员开展了微阵列（microarray）研究：在30℃条件下模拟跳蚤肠道环境，向培养基中添加3种群体感应信号，以未添加信号的同条件培养细胞作为对照。本实验使用ΔPgm（色素阴性）R88菌株，三种信号分别为1种AI-2以及2种酰基高丝氨酸内酯（AHLs），具体为N-(3-氧代辛酰基)-L-高丝氨酸内酯和N-己酰基-DL-高丝氨酸内酯。

实验总体设计如下：从添加了3种信号的培养物中提取6份独立RNA样本，与未添加信号的对照培养物的6份独立RNA样本配对，用于6张双色微阵列（two-color microarrays）的杂交实验。本研究采用染料互换（dye-swap）设计以消除Cy5与Cy3荧光染料的检测偏倚。