Abstract
Attached Vibrio cholerae biofilms are essential for environmental persistence and infectivity. The vps loci (vpsU, vpsA-K, and vpsL-Q) are required for mature biofilm formation and are responsible for the synthesis of exopolysaccharide. Transcription of vps genes is activated by the signaling molecule bis-(3′–5′)-cyclic di-GMP (c-di-GMP), whose metabolism is controlled by the proteins containing the GGDEF and/or EAL domains. The ferric uptake regulator (Fur) plays key roles in the transcription of many genes involved in iron metabolism and non-iron functions. However, roles for Fur in Vibrio biofilm production have not been documented. In this study, phenotypic assays demonstrated that Fur, independent of iron, decreases in vivo c-di-GMP levels and inhibits in vitro biofilm formation by Vibrio cholerae. The Fur box-like sequences were detected within the promoter-proximal DNA regions of vpsU, vpsA-K, vieSAB, and cdgD, suggesting that transcription of these genes may be under the direct control of Fur. Indeed, the results of luminescence, quantitative PCR (qPCR), electrophoretic mobility shift assay (EMSA), and DNase I footprinting assays demonstrated Fur to bind to the promoter-proximal DNA regions of vpsU, vpsA-K, and cdgD to repress their transcription. In contrast, Fur activates the transcription of vieSAB in a direct manner. The cdgD and vieSAB encode proteins with GGDEF and EAL domains, respectively. Thus, data presented here highlight a new physiological role for Fur wherein it acts as a repressor of V. cholerae biofilm formation mediated by decreasing the production of exopolysaccharide and the intracellular levels of c-di-GMP.
Highlights
Vibrio cholerae, an inhabitant of aquatic ecosystems, is the causative agent of the fatal waterborne diarrheal disease cholera (Clemens et al, 2017)
To characterize the biofilm formation regulated by iron and ferric uptake regulator (Fur) in V. cholerae strain C7258 (El Tor), several biofilm phenotype assays were performed, which compared biofilm morphology and quantity in the fur and wild type (WT) strains grown in normal, iron-replete, and iron-starved conditions
These results suggested that deletion of fur results in enhanced biofilm formation independent of the presence or lack of Fe2+
Summary
An inhabitant of aquatic ecosystems, is the causative agent of the fatal waterborne diarrheal disease cholera (Clemens et al, 2017). V. cholerae is most often found in the form of biofilm communities attached to biotic and abiotic surfaces (Yildiz and Visick, 2009). These biofilms have been proven essential for the environmental persistence and infectivity of V. cholerae. The expression of biofilm-related genes is upregulated within a short time, and this accounts for the immediate initiation of biofilm formation after adherence of V. cholerae to intestinal cells, and biofilm-like aggregates of V. cholerae are frequently shed in the stool of cholera patients (Faruque et al, 2006; Tamayo et al, 2010; Sengupta et al, 2016). Deletion of any of the vps genes produces smooth colony morphology with decreased biofilm formation in vitro (Fong et al, 2010, 2017)
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