title: The cyclic nucleotide monophosphate domain of Xanthomonas campestris global regulator Clp defines a new class of cyclic di-GMP effectors.
abstract: The widely conserved second messenger cyclic diguanosine monophosphate (c-di-GMP) plays a key role in quorum-sensing (QS)-dependent production of virulence factors in Xanthomonas campestris pv. campestris. The detection of QS diffusible signal factor (DSF) by the sensor RpfC leads to the activation of response regulator RpfG, which activates virulence gene expression by degrading c-di-GMP. Here, we show that a global regulator in the X. campestris pv. campestris QS regulatory pathway, Clp, is a c-di-GMP effector. c-di-GMP specifically binds to Clp with high affinity and induces allosteric conformational changes that abolish the interaction between Clp and its target gene promoter. Clp is similar to the cyclic AMP (cAMP) binding proteins Crp and Vfr and contains a conserved cyclic nucleotide monophosphate (cNMP) binding domain. Using site-directed mutagenesis, we found that the cNMP binding domain of Clp contains a glutamic acid residue (E99) that is essential for c-di-GMP binding. Substituting the residue with serine (E99S) resulted in decreased sensitivity to changes in the intracellular c-di-GMP level and attenuated bacterial virulence. These data establish the direct role of Clp in the response to fluctuating c-di-GMP levels and depict a novel mechanism by which QS links the second messenger with the X. campestris pv. campestris virulence regulon.
authors.affiliation: Institute of Molecular and Cell Biology, Singapore, Singapore.
authors.forename: Fei
authors.initials: F
authors.lastname: Tao
authors.forename: Ya-Wen
authors.initials: YW
authors.lastname: He
authors.forename: Dong-Hui
authors.initials: DH
authors.lastname: Wu
authors.forename: Sanjay
authors.initials: S
authors.lastname: Swarup
authors.forename: Lian-Hui
authors.initials: LH
authors.lastname: Zhang
mesh_terms: D000494:Allosteric Regulation; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D053038:Quorum Sensing; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D014774:Virulence; D037521:Virulence Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016959:Xanthomonas campestris / Q000737:chemistry / Q000472:pathogenicity*
chemical_list: D001426:Bacterial Proteins; C436751:Clp protein, Xanthomonas campestris; D014157:Transcription Factors; D037521:Virulence Factors; C062025:bis(3',5')-cyclic diguanylic acid; D006152:Cyclic GMP