Retracted: Dynamic complex formation between HD‐GYP, GGDEF and PilZ domain proteins regulates motility in Xanthomonas campestris

Abstract

SummaryRpfG is a member of a class of wide spread bacterial two-component regulators with an HD-GYP cyclic di-GMP phosphodiesterase domain. In the plant pathogen Xanthomonas campestris, RpfG together with the sensor kinase RpfC regulates multiple factors as a response to the cell-to-cell Diffusible Signalling Factor (DSF). A dynamic physical interaction of RpfG with two diguanylate cyclase (GGDEF) domain proteins controls motility. Here we show that, contrary to expectation, regulation of motility by the GGDEF domain proteins does not depend upon their cyclic di-GMP synthetic activity. Furthermore we show that the complex of RpfG and GGDEF domain proteins recruits a specific PilZ domain 'adaptor' protein, and this complex then interacts with the pilus motor proteins PilU and PiIT. The results support a model in which DSF signalling influences motility through the highly regulated dynamic interaction of proteins that affect pilus action. A specific motif that we identify to be required for HD-GYP domain interaction is conserved in a number of GGDEF domain proteins, suggesting that regulation via interdomain interactions is of broad relevance.

Progress in Understanding the Molecular Basis Underlying Functional Diversification of Cyclic Dinucleotide Turnover Proteins

“…In this case, the XXDXDX motif, which is highly conserved in GGDEF domains, is required for the interaction with the HD-GYP domain. HD-GYP-GGDEF complex formation serves to control motility through recruitment of a PilZ domain protein and interaction with the pilus biogenesis machinery (35,36). Overall, these data indicate that GGDEF domain proteins possess several protein interaction interfaces which participate in the formation of supramolecular complexes.…”

“…An alanine substitution within the signature HD dyad leads to a loss of both enzyme activity and regulatory action (66). In contrast, although alanine substitutions in the signature GYP motif have little or no effect on enzyme activity, they do counteract the interaction of RpfG with particular GGDEF domain proteins to modulate a specific subset of RpfG-mediated phenotypes (35,66,70).…”

“…If GGDEF domains interact directly with Y285, they need to intercalate with the inner side of the HD-GYP nucleotide binding pocket. This would prevent cyclic di-GMP binding and phosphodiesterase activity, although such effects have not been observed in vitro (35). An intriguing alternative is that RpfG involvement in protein-protein complexes is determined not only by cyclic di-GMP binding but also by conformational alterations associated with cyclic di-GMP degradation, which would be "reported" via the GYP loop.…”

“…Interactions between the EAL domain protein YciR and diguanylate cyclase YdaM control a key step in E. coli biofilm formation through a suggested modulation of localized cyclic di-GMP levels (34). Functionality is also provided, however, by specific protein-protein interactions that are independent of the catalytic activity (19,35). In this case, the XXDXDX motif, which is highly conserved in GGDEF domains, is required for the interaction with the HD-GYP domain.…”

Structural Basis of Functional Diversification of the HD-GYP Domain Revealed by the Pseudomonas aeruginosa PA4781 Protein, Which Displays an Unselective Bimetallic Binding Site

“…If this is the case, since HD-GYPs are not ubiquitous in bacteria, the presence of genes encoding these domains may well be considered a hallmark of bacterial species with more sophisticated biofilm-mediated survival strategies, such as Pseudomonas aeruginosa (9,10) or Vibrio cholerae (11). Indeed, HD-GYPs have been shown to interact with GGDEF domains within multien-zyme complexes, while EAL domains seem to interact with GGDEF domains only in hybrid proteins (i.e., when they are fused within the same polypeptide chain) (12,13). From a structural point of view, HD-GYPs are metal binding proteins named after their conserved motifs, but while the HD residues clearly serve as metal ligands, the role of the GYP triad is not so clear.…”