title: Role of (p)ppGpp in biofilm formation and expression of filamentous structures in Bordetella pertussis.
abstract: Bordetella pertussis, the causative agent of whooping cough, is highly adapted to cause human infection. The production of virulence factors, such as adhesins and toxins, is just part of an array of mechanisms by which B. pertussis causes infection. The stringent response is a global bacterial response to nutritional limitation that is mediated by the accumulation of cellular ppGpp and pppGpp [termed together as (p)ppGpp]. Here, we demonstrate that production of (p)ppGpp was controlled by RelA and SpoT proteins in B. pertussis, and that mutation-induced loss of both proteins together caused deficiencies in (p)ppGpp production. The (p)ppGpp-deficient mutants also exhibited defects in growth regulation, decreases in viability under nutritionally limited conditions, increases in susceptibility to oxidative stress and defects in biofilm formation. Analysis of the secreted proteins and the respective transcripts showed that lack of (p)ppGpp led to decreased expression of fim3 and bsp22, which encode a fimbrial subunit and the self-polymerizing type III secretion system tip protein, respectively. Moreover, electron microscopic analysis also indicated that (p)ppGpp regulated the formation of filamentous structures. Most virulence genes - including fim3 and bsp22 - were expressed in the Bvg(+) phase during which the BvgAS two-component system was activated. Although fim3 and bsp22 were downregulated in a (p)ppGpp-deficient mutant, normal expression of fhaB, cyaA and ptxA persisted. Lack of coherence between virulence gene expression and (p)ppGpp production indicated that (p)ppGpp did not modulate the Bvg phase. Taken together, our data indicate that (p)ppGpp may govern an as-yet-unrecognized system that influences B. pertussis pathogenicity.
authors.affiliation: Department of Infectious Diseases, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Sugisaki
authors.affiliation: Department of Infectious Diseases, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Hanawa
authors.affiliation: Department of Infectious Diseases, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Hideo
authors.initials: H
authors.lastname: Yonezawa
authors.affiliation: Department of Infectious Diseases, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Osaki
authors.affiliation: Department of Pharmacology, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Fukutomi
authors.affiliation: Department of Anatomy, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Hayato
authors.initials: H
authors.lastname: Kawakami
authors.affiliation: Department of Microbiology and Molecular Genetics, Graduate School of Pharmaceutical Sciences, Chiba University, 1-8-1, Inohana, Chuo-ku, Chiba-shi, Chiba 260-8675, Japan.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Yamamoto
authors.affiliation: Department of Infectious Diseases, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo 181-8611, Japan.
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kamiya
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D018441:Biofilms / Q000254:growth & development*; D001886:Bordetella pertussis / Q000235:genetics / Q000472:pathogenicity*; D010861:Fimbriae, Bacterial / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D006159:Guanosine Tetraphosphate / Q000378:metabolism*; D006801:Humans; D008025:Ligases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D011755:Pyrophosphatases / Q000235:genetics / Q000378:metabolism; D014774:Virulence; D037521:Virulence Factors
chemical_list: D001426:Bacterial Proteins; D037521:Virulence Factors; D006159:Guanosine Tetraphosphate; C027627:guanosine-3',5'-bis(diphosphate) 3'-pyrophosphatase; D011755:Pyrophosphatases; D008025:Ligases; C028812:guanosine 3',5'-polyphosphate synthetases