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

Shi-qi An, Delphine L. Caly, Yvonne McCarthy, Sarah L. Murdoch, Joseph Ward, Melanie Febrer, J. Maxwell Dow, Robert P. Ryan (Lead / Corresponding author)

    Research output: Contribution to journalArticle

    17 Citations (Scopus)
    96 Downloads (Pure)

    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 (Kd~2 µM). 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.
    Original languageEnglish
    Article numbere1004429
    Number of pages12
    JournalPLoS Pathogens
    Volume10
    Issue number10
    DOIs
    Publication statusPublished - 16 Oct 2014

    Fingerprint

    Virulence
    Proteins
    Stenotrophomonas maltophilia
    Biofilms
    Xanthomonas campestris
    bis(3',5')-cyclic diguanylic acid
    Calorimetry
    Mutation
    Second Messenger Systems
    Transcriptome
    Pseudomonas aeruginosa
    Transcription Factors
    Yeasts
    Genes

    Cite this

    An, Shi-qi ; Caly, Delphine L. ; McCarthy, Yvonne ; Murdoch, Sarah L. ; Ward, Joseph ; Febrer, Melanie ; Dow, J. Maxwell ; Ryan, Robert P. / Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence. In: PLoS Pathogens. 2014 ; Vol. 10, No. 10.
    @article{44b2d89847f64145b3346503b7fb77d8,
    title = "Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence",
    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 (Kd~2 µM). 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.",
    author = "Shi-qi An and Caly, {Delphine L.} and Yvonne McCarthy and Murdoch, {Sarah L.} and Joseph Ward and Melanie Febrer and Dow, {J. Maxwell} and Ryan, {Robert P.}",
    year = "2014",
    month = "10",
    day = "16",
    doi = "10.1371/journal.ppat.1004429",
    language = "English",
    volume = "10",
    journal = "PLoS Pathogens",
    issn = "1553-7366",
    publisher = "Public Library of Science",
    number = "10",

    }

    An, S, Caly, DL, McCarthy, Y, Murdoch, SL, Ward, J, Febrer, M, Dow, JM & Ryan, RP 2014, 'Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence', PLoS Pathogens, vol. 10, no. 10, e1004429. https://doi.org/10.1371/journal.ppat.1004429

    Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence. / An, Shi-qi; Caly, Delphine L.; McCarthy, Yvonne; Murdoch, Sarah L.; Ward, Joseph; Febrer, Melanie; Dow, J. Maxwell; Ryan, Robert P. (Lead / Corresponding author).

    In: PLoS Pathogens, Vol. 10, No. 10, e1004429, 16.10.2014.

    Research output: Contribution to journalArticle

    TY - JOUR

    T1 - Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence

    AU - An, Shi-qi

    AU - Caly, Delphine L.

    AU - McCarthy, Yvonne

    AU - Murdoch, Sarah L.

    AU - Ward, Joseph

    AU - Febrer, Melanie

    AU - Dow, J. Maxwell

    AU - Ryan, Robert P.

    PY - 2014/10/16

    Y1 - 2014/10/16

    N2 - 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 (Kd~2 µM). 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.

    AB - 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 (Kd~2 µM). 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.

    U2 - 10.1371/journal.ppat.1004429

    DO - 10.1371/journal.ppat.1004429

    M3 - Article

    VL - 10

    JO - PLoS Pathogens

    JF - PLoS Pathogens

    SN - 1553-7366

    IS - 10

    M1 - e1004429

    ER -