Cell-cell signal-dependent dynamic interactions between HD-GYP and GGDEF domain proteins mediate virulence in Xanthomonas campestris.
RpfG is a paradigm for a class of widespread bacterial two-component regulators with a CheY-like receiver domain attached to a histidine-aspartic acid-glycine-tyrosine-proline (HD-GYP) cyclic di-GMP phosphodiesterase domain. In the plant pathogen Xanthomonas campestris pv. campestris (Xcc), a two-component system comprising RpfG and the complex sensor kinase RpfC is implicated in sensing and responding to the diffusible signaling factor (DSF), which is essential for cell-cell signaling. RpfF is involved in synthesizing DSF, and mutations of rpfF, rpfG, or rpfC lead to a coordinate reduction in the synthesis of virulence factors such as extracellular enzymes, biofilm structure, and motility. Using yeast two-hybrid analysis and fluorescence resonance energy transfer experiments in Xcc, we show that the physical interaction of RpfG with two proteins with diguanylate cyclase (GGDEF) domains controls a subset of RpfG-regulated virulence functions. RpfG interactions were abolished by alanine substitutions of the three residues of the conserved GYP motif in the HD-GYP domain. Changing the GYP motif or deletion of the two GGDEF-domain proteins reduced Xcc motility but not the synthesis of extracellular enzymes or biofilm formation. RpfG-GGDEF interactions are dynamic and depend on DSF signaling, being reduced in the rpfF mutant but restored by DSF addition. The results are consistent with a model in which DSF signal transduction controlling motility depends on a highly regulated, dynamic interaction of proteins that influence the localized expression of cyclic di-GMP.
Department of Microbiology, BIOMERIT Research Centre, BioSciences Institute, University College Cork, Cork, Ireland.
Robert P
RP
Ryan
Yvonne
Y
McCarthy
Maxuel
M
Andrade
Chuck S
CS
Farah
Judith P
JP
Armitage
J Maxwell
JM
Dow
D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D031541:Fluorescence Resonance Energy Transfer; D056265:Microbial Interactions; D016297:Mutagenesis, Site-Directed; D010944:Plants / Q000382:microbiology; D054730:Protein Interaction Domains and Motifs; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques; D014774:Virulence / Q000235:genetics / Q000502:physiology; D016959:Xanthomonas campestris / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; C419552:RpfG protein, Xanthomonas campestris