Quorum sensing control of phosphorus acquisition in Trichodesmium consortia

Abstract

Colonies of the cyanobacterium Trichodesmium are abundant in the oligotrophic ocean, and through their ability to fix both CO(2) and N(2), have pivotal roles in the cycling of carbon and nitrogen in these highly nutrient-depleted environments. Trichodesmium colonies host complex consortia of epibiotic heterotrophic bacteria, and yet, the regulation of nutrient acquisition by these epibionts is poorly understood. We present evidence that epibiotic bacteria in Trichodesmium consortia use quorum sensing (QS) to regulate the activity of alkaline phosphatases (APases), enzymes used by epibionts in the acquisition of phosphate from dissolved-organic phosphorus molecules. A class of QS molecules, acylated homoserine lactones (AHLs), were produced by cultivated epibionts, and adding these AHLs to wild Trichodesmium colonies collected at sea led to a consistent doubling of APase activity. By contrast, amendments of (S)-4,5-dihydroxy-2,3-pentanedione (DPD)-the precursor to the autoinducer-2 (AI-2) family of universal interspecies signaling molecules-led to the attenuation of APase activity. In addition, colonies collected at sea were found by high performance liquid chromatography/mass spectrometry to contain both AHLs and AI-2. Both types of molecules turned over rapidly, an observation we ascribe to quorum quenching. Our results reveal a complex chemical interplay among epibionts using AHLs and AI-2 to control access to phosphate in dissolved-organic phosphorus.

Figure 1

APase activity of Trichodesmium colonies in response to amendment with a cocktail of saturated, long-chain AHLs (C10-HSL, C12-HSL and C14-HSL) in the North Atlantic and North Pacific. Error bars indicate the magnitude of the range for triplicate incubations. As the assays were conducted with slightly different methods in the North Atlantic and North Pacific, data are presented as ratios to the no amendment control incubations. The horizontal dashed line represents no response to the amendments.

Figure 2

Representative photomicrographs of Trichodesmium colonies from the North Atlantic showing the cell-specific response to an enzyme labeled fluorescence assay of APase activity. The bright white and green areas on or near the orange autofluorescent Trichodesmium trichomes indicate localized APase activity. Left: endogenous Trichodesmium APase activity from a colony that did not receive an AHL amendment. Right: a colony from an incubation amended with the AHL cocktail, which shows the APase activity of epibiotic bacteria.

Figure 3

APase activity of Trichodesmium colonies in response to various AHL and AI-2 amendments (shown in legend) at a station in the North Pacific Ocean. Error bars indicate the magnitude of the range for triplicate incubations. Data are presented as ratios to the no amendment control incubations. The horizontal dashed line represents no response to the amendments.

Figure 4

AHL and AI-2 in Trichodesmium colonies and amendment experiments. (a) HPLC/MS/MS chromatograms showing the presence of C14-HSL in Trichodesmium colonies. The top chromatogram shows the selected reaction monitoring (SRM) transition from the parent ion of C14-HSL to the acyl chain ion, whereas the bottom chromatogram shows the transition to the lactone ring ion. (b) Chromatogram showing the SRM transition from the parent ion of tagged DPD to a selective fragment ion. Scheme for the tagging of AI-2 is also shown. Tagging is necessary to both stabilize the molecule and make AI-2 amenable to HPLC/MS/MS detection (Campagna et al., 2009). (c and d) Plots showing the concentrations of C8-HSL and AI-2 through time in incubations containing Trichodesmium colonies. Amendments are shown in the legend. Error bars indicate the magnitude of the range for triplicate incubations.