The putative sensor histidine kinase PhcK is required for the full expression ofphcAencoding the global transcriptional regulator to drive the quorum-sensing circuit ofRalstonia solanacearumstrain OE1-1

A gram-negative plant-pathogenic bacteriumRalstonia solanacearumstrain 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 ofphcAregulation 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 aphcK-deletion mutant (Delta phcK) that produced significantly less EPS I and ralfuranone than the wild-type strain OE1-1. Quantitative reverse transcription PCR assays showed that thephcAexpression level was significantly down-regulated in the Delta phcKmutant 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 Delta phcKmutant, whereas the expression levels of 85.9% of the PhcA-negatively regulated genes were up-regulated. Additionally, the nativephcK-expressing complemented Delta phcKstrain and the Delta phcKmutant transformed withphcAcontrolled by a constitutive promoter recovered their cell aggregation phenotypes. Considered together, the results of this study indicate thatphcKis required for fullphcAexpression, thereby driving the QS circuit ofR. solanacearumstrain OE1-1. This is the first report of thephcAtranscriptional regulation ofR. solanacearum.