RETRACTED: Cell–cell signaling in Xanthomonas campestris involves an HD-GYP domain protein that functions in cyclic di-GMP turnover

Abstract

HD-GYP is a protein domain of unknown biochemical function implicated in bacterial signaling and regulation. In the plant pathogen Xanthomonas campestris pv. campestris , the synthesis of virulence factors and dispersal of biofilms are positively controlled by a two-component signal transduction system comprising the HD-GYP domain regulatory protein RpfG and cognate sensor RpfC and by cell–cell signaling mediated by the diffusible signal molecule DSF (diffusible signal factor). The RpfG/RpfC two-component system has been implicated in DSF perception and signal transduction. Here we show that the role of RpfG is to degrade the unusual nucleotide cyclic di-GMP, an activity associated with the HD-GYP domain. Mutation of the conserved H and D residues of the isolated HD-GYP domain resulted in loss of both the enzymatic activity against cyclic di-GMP and the regulatory activity in virulence factor synthesis. Two other protein domains, GGDEF and EAL, are already implicated in the synthesis and degradation respectively of cyclic di-GMP. As with GGDEF and EAL domains, the HD-GYP domain is widely distributed in free-living bacteria and occurs in plant and animal pathogens, as well as beneficial symbionts and organisms associated with a range of environmental niches. Identification of the role of the HD-GYP domain thus increases our understanding of a signaling network whose importance to the lifestyle of diverse bacteria is now emerging.

Cell–cell signal-dependent dynamic interactions between HD-GYP and GGDEF domain proteins mediate virulence in Xanthomonas campestris

“…The role of protein-protein interactions involving RpfG in the regulation of virulence factor synthesis was examined by assessing in parallel the effects of alanine substitution of the residues of the GYP motif within RpfG and the effects of the mutation of the genes encoding the GGDEF target proteins XC_0249 and XC_0420. We previously have shown that expression of the HD-GYP domain of RpfG in the absence of the REC domain can restore the synthesis of virulence factors and motility in the rpfG mutant of Xcc to wild-type levels (6). These diverse regulatory effects are prevented by alanine substitution in the HD dyad (Fig.…”

“…campestris (Xcc) has been shown to be the unsaturated fatty acid cis-11-methyl-dodecenoic acid (5). Both the synthesis and perception of the DSF signal require products of the regulation of pathogenicity factors (rpf) gene cluster (1,4,6). The synthesis of DSF is dependent on RpfF, which has some amino acid sequence similarity to enoyl CoA hydratases, whereas the two-component system comprising the sensor kinase RpfC and regulator RpfG is implicated in DSF perception and signal transduction (1,4,6,7).…”

“…RpfC is a complex sensor kinase with a predicted membraneassociated sensory input domain. RpfG is a regulator with a CheY-like receiver (REC) domain attached to a histidine-aspartic acid-glycine-tyrosine-proline (HD-GYP) domain which acts to degrade the second messenger, bis (3′,5′)-cyclic diguanosine monophosphate (cyclic di-GMP) (6). Mutation of rpfF, rpfG, and rpfC in Xcc leads to a coordinate reduction in the synthesis of virulence factors, including extracellular protease, endoglucanase, and endomannanase enzymes and the extracellular polysaccharide xanthan; alterations in biofilm formation; reduced pilusdependent motility; and a reduction in virulence (1,4,7,8).…”

Genetic reductionist approach for dissecting individual roles of GGDEF proteins within the c-di-GMP signaling network in Salmonella

“…The only known effector domain that synthesizes c-di-GMP (designated as diguanylate cyclase activity, DGC) is the GGDEF domain. Two domains, EAL and HD-GYP, hydrolyze c-di-GMP (designated as phosphodiesterase activity, PDE) into linear 5Ј-phosphoguanylyl-(3Ј-5Ј)-guanosine (pGpG) or guanosine monophosphate (GMP), respectively (7)(8)(9)(10)(11)(12)(13). Many DGC proteins also contain an RxxD conserved motif 5 amino acids upstream from the GGDEF motif.…”

Diguanylate cyclase activity of the Mycobacterium leprae T cell antigen ML1419c

“…Approximately 50% of proteins with characterized DGC activity (Römling et al, 2013) are regulated via allosteric control and the binding of cyclic di-GMP to a conserved inhibitory site (I-site) motif (RxxD) directly upstream of the GGDEF motif (Christen et al, 2006;Wassmann et al, 2007). Phosphodiesterase (PDE) proteins that possess EAL or HD-GYP domains are responsible for the depletion of cyclic di-GMP and conversion into 5¢-phosphoguanylyl-(3¢,5¢)-guanosine (pGpG) or two molecules of guanosine monophosphate (GMP), respectively (Ryan et al, 2006;Schmidt et al, 2005). Recently, oligoribonuclease (Orn) has also been identified to participate in the conversion of pGpG into GMP in Pseudomonas aeruginosa (Cohen et al, 2015;Orr et al, 2015).…”