Fig. 2: d-Arg production and chemotactic response are coordinated by RpoS.
From: d-amino acids signal a stress-dependent run-away response in Vibrio cholerae
a, Left: localization of sfGFP-MCPDRK expressed from its native locus and promoter at exponential and stationary phase. Representative micrographs of 3 independent replicates are shown; at least 3 images were acquired per timepoint. Scale bar, 2 μm. Right: demographic analysis of the fluorescence intensity of sfGFP-MCPDRK relative to cell length (n > 500 cells). The colour bar represents fluorescence intensity. b, Bottom: growth-phase dependent subcellular localization of sfGFP-MCPDRK. Filled grey circles, OD600  of V. cholerae grown in TB; open tradewind-blue circles, percentage of cells with sfGFP-MCPDRK polar foci. Top: western blot against sfGFP-MCPDRK using anti-GFP-specific antibody at the indicated OD600. Samples were normalized and loaded with equal protein amount. c, Growth curve of V. cholerae grown in MSR6 minimal medium supplemented with 0.04% w/v d-glucose and 0.4% w/v succinate (grey line) at the indicated timepoints. The corresponding percentage of cells with sfGFP-MCPDRK foci (open tradewind-blue circles) is also indicated. Red (glucose depletion) and purple (succinate depletion) arrows indicate carbon starvation. Quantification of % of cells with polar foci in b and c was based on a single experiment where >500 cells were counted from 3 different images per timepoint. d, Left: micrographs showing the expression of sfGFP-MCPDRK in a ΔrpoS mutant background. Representative micrographs of 3 independent replicates are shown; at least 3 images were acquired per timepoint: scale bar, 2 μm. Right: demographic analysis of the fluorescence intensity of sfGFP-MCPDRK relative to cell length. The colour bar represents fluorescence intensity. e, Chemotaxis response of ΔrpoS mutant to d-Arg. Black diamonds represent the mean of 3 independent biological replicates. Error bars represent mean ± s.d. of 3 biologically independent replicates. Significant differences (unpaired t-test) are indicated by ***P < 0.001. f, DAA production by the different strains. Significant differences (unpaired t-test) are indicated by **P < 0.01 or ***P < 0.001.
