Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence

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Date
2014
Authors
An, Shi-Qi
Caly, Delphine L.
McCarthy, Yvonne
Murdoch, Sarah L.
Ward, Joseph
Febrer, Melanie
Dow, J. Maxwell
Ryan, Robert P.
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Volume Title
Publisher
Public Library of Science
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Abstract
Bis-(3 ',5 ') cyclic di-guanylate (cyclic di-GMP) is a key bacterial second messenger that is implicated in the regulation of many critical processes that include motility, biofilm formation and virulence. Cyclic di-GMP influences diverse functions through interaction with a range of effectors. Our knowledge of these effectors and their different regulatory actions is far from complete, however. Here we have used an affinity pull-down assay using cyclic di-GMP-coupled magnetic beads to identify cyclic di-GMP binding proteins in the plant pathogen Xanthomonas campestris pv. campestris (Xcc). This analysis identified XC_3703, a protein of the YajQ family, as a potential cyclic di-GMP receptor. Isothermal titration calorimetry showed that the purified XC_3703 protein bound cyclic di-GMP with a high affinity (K-d similar to 2 mu M). Mutation of XC_3703 led to reduced virulence of Xcc to plants and alteration in biofilm formation. Yeast two-hybrid and far-western analyses showed that XC_3703 was able to interact with XC_2801, a transcription factor of the LysR family. Mutation of XC_2801 and XC_3703 had partially overlapping effects on the transcriptome of Xcc, and both affected virulence. Electromobility shift assays showed that XC_3703 positively affected the binding of XC_2801 to the promoters of target virulence genes, an effect that was reversed by cyclic di-GMP. Genetic and functional analysis of YajQ family members from the human pathogens Pseudomonas aeruginosa and Stenotrophomonas maltophilia showed that they also specifically bound cyclic di-GMP and contributed to virulence in model systems. The findings thus identify a new class of cyclic di-GMP effector that regulates bacterial virulence.
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Keywords
PilZ domain proteins , Campestris pv. campestris , Diffusible signal factor , Airway epithelial cells , Cystic fibrosis airway , Xanthomonas campestris , Pseudomonas aeruginosa , HD-GYP , Biofilm formation , 2nd messenger
Citation
An S-q, Caly DL, McCarthy Y, Murdoch SL, Ward J, Febrer M, et al. (2014) Novel Cyclic di-GMP Effectors of the YajQ Protein Family Control Bacterial Virulence. PLoS Pathog 10(10): e1004429. doi:10.1371/journal.ppat.1004429