Fig. 1: d-Arg and d-Lys signal a chemorepellent response in V. cholerae through the putative MCP VC1313.
From: d-amino acids signal a stress-dependent run-away response in Vibrio cholerae
a, Schematic representation of DAA racemization. The most abundant amino acids produced are shown (open circles, l-amino acids; filled circles, d-amino acids). OM, outer membrane; IM, inner membrane; CYT, cytoplasm. b, Representative images of V. cholerae wild type (wt) and ΔbsrV mutant expanding in 0.3% soft agar. Images are representative of experiments repeated at least three times. c, Motility and d-amino acid production analysis of BsrV derivatives; grey bars indicate relative motility compared to wt or ΔbsrV knockout mutant, catalytic site mutant (BsrV K95A) and the complemented strain harbouring bsrV under an aTc inducible promoter (pbsrV); green dots represent the amount of secreted d-amino acids. pĂ˜, empty plasmid. Error bars represent mean ± s.d. of 3 biologically independent replicates. Significant differences (paired t-test) are indicated by **P < 0.01 or ***P < 0.001. d, Motility of ΔbsrV mutant in soft-agar plates that are chemically complemented with 5 mM d-amino acids are shown relative to the wild-type strain. Error bars represent mean ± s.d. of 6 biologically independent replicates examined over 2 independent experiments. Significant differences (one-way analysis of variance (ANOVA)) are indicated by ****P < 0.0001. e, Chemotactic response to 0.1 mM d-amino acids in capillary assays. Chemotaxis ratio (CR) was calculated relative to the control capillary containing no stimulus. CR > 1, attractants; CR < 1, repellents; CR = 1, no response. Black diamonds represent the mean of 3 independent biological replicates. Significant differences (unpaired t-test) are indicated by *P < 0.05 or ***P < 0.001. f, Thermal proteome profiling analysis of V. cholerae cell extracts exposed to d-Arg. Stabilized and destabilized proteins in the presence of d-Arg are highlighted in red and green, respectively. From the 45 MCPs found in V. cholerae’s genome (black), only 3 interacted with d-Arg: VC2161, VC1313 and VC1406. g, Chemotactic response of selected MCP candidates to d-Arg. Double mutants (ΔbsrV Δmcp) were constructed and the ability to respond to d-Arg was tested by capillary assays. ΔbsrV strain was used as background. LAA, l-amino acids; DAA, d-amino acids. Black diamonds represent the mean of 3 independent biological replicates. Significant differences (unpaired t-test) are indicated by ***P < 0.001.
