title: The exopolysaccharide gene cluster Bcam1330-Bcam1341 is involved in Burkholderia cenocepacia biofilm formation, and its expression is regulated by c-di-GMP and Bcam1349.
abstract: In Burkholderia cenocepacia, the second messenger cyclic diguanosine monophosphate (c-di-GMP) has previously been shown to positively regulate biofilm formation and the expression of cellulose and type-I fimbriae genes through binding to the transcriptional regulator Bcam1349. Here, we provide evidence that cellulose and type-I fimbriae are not involved in B. cenocepacia biofilm formation in flow chambers, and we identify a novel Bcam1349/c-di-GMP-regulated exopolysaccharide gene cluster which is essential for B. cenocepacia biofilm formation. Overproduction of Bcam1349 in trans promotes wrinkly colony morphology, pellicle, and biofilm formation in B. cenocepacia. A screen for transposon mutants unable to respond to the overproduction of Bcam1349 led to the identification of a 12-gene cluster, Bcam1330-Bcam1341, the products of which appear to be involved in the production of a putative biofilm matrix exopolysaccharide and to be essential for flow-chamber biofilm formation. We demonstrate that Bcam1349 binds to the promoter region of genes in the Bcam1330-Bcam1341 cluster and that this binding is enhanced by the presence of c-di-GMP. Furthermore, we demonstrate that overproduction of both c-di-GMP and Bcam1349 leads to increased transcription of these genes, indicating that c-di-GMP and Bcam1349 functions together in regulating exopolysaccharide production from the Bcam1330-Bcam1341 gene cluster. Our results suggest that the product encoded by the Bcam1330-Bcam1341 gene cluster is a major exopolysaccharide that provides structural stability to the biofilms formed by B. cenocepacia, and that its production is regulated by c-di-GMP through binding to and promotion of the activity of the transcriptional regulator Bcam1349.
authors.affiliation: Department of International Health, Immunology and Microbiology, Faculty of Health Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Mustafa
authors.initials: M
authors.lastname: Fazli
authors.forename: Yvonne
authors.initials: Y
authors.lastname: McCarthy
authors.forename: Michael
authors.initials: M
authors.lastname: Givskov
authors.forename: Robert P
authors.initials: RP
authors.lastname: Ryan
authors.forename: Tim
authors.initials: T
authors.lastname: Tolker-Nielsen
mesh_terms: D018441:Biofilms / Q000254:growth & development*; D053898:Biosynthetic Pathways / Q000235:genetics*; D057508:Burkholderia cenocepacia / Q000235:genetics / Q000502:physiology*; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000378:metabolism; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000378:metabolism; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D005810:Multigene Family; D016254:Mutagenesis, Insertional; D011135:Polysaccharides, Bacterial / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014157:Transcription Factors / Q000378:metabolism*
chemical_list: D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D011135:Polysaccharides, Bacterial; D014157:Transcription Factors; C062025:bis(3',5')-cyclic diguanylic acid; D006152:Cyclic GMP