Laboratory of Microbial Interactions, Department of Molecular and Cellular Interactions (VIB), Vrije Universiteit Brussel (VUB), Pleinlaan 2, B-1050 Brussels, Belgium.
Microbiology (Reading). 2010 Mar;156(Pt 3):678-686. doi: 10.1099/mic.0.031971-0. Epub 2009 Nov 19.
The LysR-type transcriptional regulator (LTTR) OxyR orchestrates the defence of the opportunistic pathogen Pseudomonas aeruginosa against reactive oxygen species. In previous work we also demonstrated that OxyR is needed for the utilization of the ferrisiderophore pyoverdine, stressing the importance of this regulator. Here, we show that an oxyR mutant is unable to swarm on agar plates, probably as a consequence of absence of production of rhamnolipid surfactant molecules. Another obvious phenotypic change was the increased production of the phenazine redox-active molecule pyocyanin in the oxyR mutant. As already described, the oxyR mutant could not grow in LB medium, unless high numbers of cells (>10( 8) ml(-1)) were inoculated. However, its growth in Pseudomonas P agar (King's A), a medium inducing pyocyanin production, was like that of the wild-type, suggesting a protective action of this redox-active phenazine compound. This was confirmed by the restoration of the capacity to grow in LB medium upon addition of pure pyocyanin. Although both rhamnolipid and pyocyanin production are controlled by quorum sensing, no obvious changes were observed in the production of N-acylhomoserine lactones or the Pseudomonas quinolone signal (PQS). Complementation of rhamnolipid production and motility, and restoration of normal pyocyanin levels, was only possible when the oxyR gene was in single copy, while pyocyanin levels were increased when oxyR was present in a multicopy vector. Conversely, plating efficiency was increased only when the oxyR gene was present in multicopy, but not when in single copy in the chromosome, due to lower expression of oxyR compared with the wild-type, suggesting that some phenotypes are differently affected in function to the levels of OxyR molecules in the cell. Analysis of transcripts of oxidative stress-response enzymes showed a strong decrease of katB, ahpC and ahpB expression in the oxyR mutant grown in LB, but this was not the case when the mutant was grown on P agar, suggesting that the OxyR dependency for the transcription of these genes is not total.
LysR 型转录调节因子（LTTR）OxyR 协调机会性病原体铜绿假单胞菌对活性氧的防御。在之前的工作中，我们还证明了 OxyR 对于利用铁载体吡咯并喹啉啉（pyoverdine）是必需的，这凸显了该调节剂的重要性。在这里，我们表明 oxyR 突变体无法在琼脂平板上群集，这可能是由于缺乏鼠李糖脂表面活性剂分子的产生所致。另一个明显的表型变化是 oxyR 突变体中吩嗪类氧化还原活性分子吡咯并喹啉啉（pyocyanin）的产量增加。如前所述，oxyR 突变体不能在 LB 培养基中生长，除非接种大量细胞（>10（8）ml（-1））。然而，它在 Pseudomonas P 琼脂（King's A）中的生长情况与野生型相似，这表明这种氧化还原活性吩嗪化合物具有保护作用。这通过添加纯吡咯并喹啉啉恢复在 LB 培养基中生长的能力得到了证实。尽管鼠李糖脂和吡咯并喹啉啉的产生都受到群体感应的控制，但 N-酰基高丝氨酸内酯或假单胞菌喹诺酮信号（PQS）的产生没有明显变化。只有当 oxyR 基因是单拷贝时，才能补充鼠李糖脂的产生和运动，并恢复正常的吡咯并喹啉啉水平，而当 oxyR 存在于多拷贝载体中时，吡咯并喹啉啉水平会增加。相反，只有当 oxyR 基因存在于多拷贝中时，平板效率才会增加，而当 oxyR 基因存在于染色体的单拷贝中时则不会增加，这是由于与野生型相比，oxyR 的表达较低，表明某些表型在功能上受到细胞中 OxyR 分子水平的不同影响。对氧化应激反应酶的转录物的分析表明，在 LB 中生长的 oxyR 突变体中 katB、ahpC 和 ahpB 的表达强烈下降，但在 P 琼脂上生长时则不是这种情况，这表明这些基因的转录对 OxyR 的依赖性不是完全的。
