Ulrich Ricky L, Hines Harry B, Parthasarathy N, Jeddeloh Jeffrey A
Bacteriology Division, United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, Maryland 21702-5011, USA.
J Bacteriol. 2004 Jul;186(13):4350-60. doi: 10.1128/JB.186.13.4350-4360.2004.
Numerous gram-negative bacteria communicate and regulate gene expression through a cell density-responsive mechanism termed quorum sensing (QS), which involves the synthesis and perception of diffusible N-acyl-homoserine lactones (AHL). In this study we genetically and physiologically characterized the Burkholderia thailandensis DW503 QS network. In silico analysis of the B. thailandensis genome revealed the presence of at least three AHL synthases (AHS) and five transcriptional regulators belonging to the LuxIR family of proteins. Mass spectrometry demonstrated that wild-type B. thailandensis synthesizes N-hexanoyl-homoserine lactone (C6-HSL), N-octanoyl-homoserine lactone (C8-HSL), and N-decanoyl-homoserine lactone (C10-HSL). Mutation of the btaI1 (luxI) AHS gene prevented accumulation of C8-HSL in culture supernatants, enhanced beta-hemolysis of sheep erythrocytes, increased lipase production, and altered colony morphology on swarming and twitching motility plates. Disruption of the btaI3 (luxI) AHS prevented biosynthesis of C6-HSL and increased lipase production and beta-hemolysis, whereas mutagenesis of the btaI2 (luxI) allele eliminated C10-HSL accumulation and reduced lipase production. Complementation of the btaI1 and btaI3 mutants fully restored the synthesis of C8-HSL and C6-HSL to parental levels. In contrast, mutagenesis of the btaR1, btaR3, btaR4, and btaR5 (luxR) transcriptional regulators had no effect on AHL accumulation, enhanced lipase production, and resulted in extensive beta-hemolysis on sheep blood agar plates. Furthermore, interruption of the btaI1, btaR1, and btaR3 genes altered colony morphology on twitching and swarming motility plates and induced pigmentation. Additionally, phenotypic microarray analysis indicated that QS in B. thailandensis both positively and negatively affects the metabolism of numerous substrates, including citric acid, formic acid, glucose 6-phosphate, capric acid, gamma-hydroxybutyric acid, and d-arabinose. These results demonstrate that mutagenesis of the B. thailandensis QS system affects various cellular processes, including lipase production, swarming and twitching motility, beta-hemolysis of sheep erythrocytes, and carbon metabolism and/or transport.
许多革兰氏阴性菌通过一种称为群体感应（QS）的细胞密度响应机制进行通讯并调节基因表达，该机制涉及可扩散的N-酰基高丝氨酸内酯（AHL）的合成与感知。在本研究中，我们对泰国伯克霍尔德菌DW503的QS网络进行了遗传学和生理学特征分析。对泰国伯克霍尔德菌基因组的计算机分析表明，其存在至少三种AHL合成酶（AHS）和五种属于LuxIR蛋白家族的转录调节因子。质谱分析表明，野生型泰国伯克霍尔德菌可合成N-己酰高丝氨酸内酯（C6-HSL）、N-辛酰高丝氨酸内酯（C8-HSL）和N-癸酰高丝氨酸内酯（C10-HSL）。btaI1（luxI）AHS基因突变可阻止C8-HSL在培养上清液中的积累，增强绵羊红细胞的β-溶血作用，增加脂肪酶产量，并改变在群体运动和颤动运动平板上的菌落形态。btaI3（luxI）AHS基因的破坏可阻止C6-HSL的生物合成，并增加脂肪酶产量和β-溶血作用，而btaI2（luxI）等位基因突变可消除C10-HSL的积累并降低脂肪酶产量。btaI1和btaI3突变体的互补作用可将C8-HSL和C6-HSL的合成完全恢复到亲本水平。相比之下，btaR1、btaR3、btaR4和btaR5（luxR）转录调节因子的突变对AHL积累没有影响，但增强了脂肪酶产量，并导致在绵羊血琼脂平板上出现广泛的β-溶血现象。此外，btaI1、btaR1和btaR3基因的中断改变了在颤动和群体运动平板上的菌落形态并诱导色素沉着。此外，表型微阵列分析表明，泰国伯克霍尔德菌中的QS对包括柠檬酸、甲酸、6-磷酸葡萄糖、癸酸、γ-羟基丁酸和D-阿拉伯糖在内的多种底物的代谢具有正向和负向影响。这些结果表明，泰国伯克霍尔德菌QS系统的突变会影响各种细胞过程，包括脂肪酶产生、群体运动和颤动运动、绵羊红细胞的β-溶血作用以及碳代谢和/或运输。
