A gram‐negative plant‐pathogenic bacterium Ralstonia solanacearum strain OE1‐1 produces and extracellularly secretes methyl 3‐hydroxymyristate (3‐OH MAME), and senses the chemical as a quorum‐sensing (QS) signal, activating QS. During QS a functional global transcriptional regulator PhcA, through the 3‐OH MAME‐dependent two‐component system, induces the production of virulence factors including a major extracellular polysaccharide EPS I and ralfuranone. To elucidate the mechanisms of phcA regulation underlying the QS system, among Tn5‐mutants from the strain OE1‐1, we identified a mutant of RSc1351 gene (phcK), encoding a putative sensor histidine kinase, that exhibited significantly decreased QS‐dependent cell aggregation. We generated a phcK‐deletion mutant (ΔphcK) that produced significantly less EPS I and ralfuranone than the wild‐type strain OE1‐1. Quantitative reverse transcription PCR assays showed that the phcA expression level was significantly down‐regulated in the ΔphcK mutant but not in other QS mutants. The transcriptome data generated with RNA sequencing technology revealed that the expression levels of 88.2% of the PhcA‐positively regulated genes were down‐regulated in the ΔphcK mutant, whereas the expression levels of 85.9% of the PhcA‐negatively regulated genes were up‐regulated. Additionally, the native phcK‐expressing complemented ΔphcK strain and the ΔphcK mutant transformed with phcA controlled by a constitutive promoter recovered their cell aggregation phenotypes. Considered together, the results of this study indicate that phcK is required for full phcA expression, thereby driving the QS circuit of R. solanacearum strain OE1‐1. This is the first report of the phcA transcriptional regulation of R. solanacearum.

中文翻译：

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假定的传感器组氨酸激酶PhcK是phcA的完整表达的必需蛋白，其编码全球转录调节因子以驱动青枯雷尔氏菌菌株OE1-1的群体感应电路。

革兰氏阴性植物致病性青枯雷尔氏菌菌株OE1-1产生并在细胞外分泌3-羟基肉豆蔻酸甲酯（3-OH MAME），并将该化学物质感应为群体感应（QS）信号，从而激活QS。在QS期间，功能性的全局转录调节剂PhcA通过3-OH MAME依赖性的双组分系统诱导产生毒力因子，包括主要的细胞外多糖EPS I和ralfuranone。为了阐明QS系统基础的phcA调控机制，在OE1-1菌株的Tn 5突变体中，我们鉴定了一个RSc1351基因突变体（phcK），编码一个假定的传感器组氨酸激酶，表现出显着降低的QS依赖性细胞聚集。我们生成了一个phcK缺失突变体（ΔphcK），与野生型OE1-1相比，该突变体产生的EPS I和ralfuranone明显减少。定量逆转录PCR分析表明，在ΔphcK突变体中，phcA表达水平显着下调，而在其他QS突变体中则没有。与RNA测序技术产生的转录组数据显示的的pHCA-正调控基因的88.2％的表达水平的Δ个下调phcK突变体，而PhcA负调控基因的表达水平为85.9％。此外，天然phcK -表达补充Δ phcK菌株和Δ phcK突变体转化的pHCA由组成型启动子控制的恢复了它们的细胞聚集的表型。综合考虑，这项研究的结果表明phcK是完整phcA表达所必需的，从而驱动了茄形青霉菌株OE1-1的QS回路。这是关于青枯菌phcA转录调控的首次报告。