Transcriptome analysis of the Vibrio fischeri LuxR-LuxI regulon

Abstract

The Vibrio fischeri quorum-sensing signal N-3-oxohexanoyl-l-homoserine lactone (3OC6-HSL) activates expression of the seven-gene luminescence operon. We used microarrays to unveil 18 additional 3OC6-HSL-controlled genes, 3 of which had been identified by other means previously. We show most of these genes are regulated by the 3OC6-HSL-responsive transcriptional regulator LuxR directly. This demonstrates that V. fischeri quorum sensing regulates a substantial number of genes other than those involved in light production.

FIG. 1.

Genetic organization and transcription activation of 3OC6-HSL-controlled genes. White arrows represent 3OC6-HSL-activated genes, and a black arrow represents the 3OC6-HSL-repressed gene. Genes previously reported as 3OC6-HSL controlled are represented by gray arrows. The directions of the arrows indicate the DNA strand. The values for gene activation (relative units of GFP fluorescence per OD unit) are shown above the first gene in each operon and are from the E. coli expression studies described in the text. In all cases, basal levels of expression with LuxR and without 3OC6-HSL or without LuxR and with 3OC6-HSL were <200 fluorescence units. The values are means of three independent experiments, and the ranges were ±28% of the means.

FIG. 2.

Gel shift assays showing direct binding of purified LuxR to promoters of 3OC6-HSL-regulated genes in vitro. Experiments were performed as previously described (28, 37). Reaction mixtures contained 1 fmol of DNA, 6 μM 3OC6-HSL, and either no LuxR (−) or LuxR at a final concentration of 6.2 nM (+). DNA probes were generated by PCR amplification of promoter regions using the transcriptional fusion plasmids as templates. Probes are shown side by side to facilitate comparison, even though sizes are different. VF0090 is shown as an example of a promoter fragment that is not shifted by LuxR, and this serves as a negative control.