Abstract
Secreted exopolysaccharides present important determinants for bacterial biofilm formation, survival, and virulence. Cellulose secretion typically requires the concerted action of a c-di-GMP-responsive inner membrane synthase (BcsA), an accessory membrane-anchored protein (BcsB), and several additional Bcs components. Although the BcsAB catalytic duo has been studied in great detail, its interplay with co-expressed subunits remains enigmatic. Here we show that E. coli Bcs proteins partake in a complex protein interaction network. Electron microscopy reveals a stable, megadalton-sized macromolecular assembly, which encompasses most of the inner membrane and cytosolic Bcs components and features a previously unobserved asymmetric architecture. Heterologous reconstitution and mutational analyses point toward a structure–function model, where accessory proteins regulate secretion by affecting both the assembly and stability of the system. Altogether, these results lay the foundation for more comprehensive models of synthase-dependent exopolysaccharide secretion in biofilms and add a sophisticated secretory nanomachine to the diverse bacterial arsenal for virulence and adaptation.
Highlights
Secreted exopolysaccharides present important determinants for bacterial biofilm formation, survival, and virulence
E. colilike bcs operons are widespread among β-Proteobacteria and γProteobacteria, including pathogenic and biocontrol organisms such as Salmonella enterica serovar Typhimurium, Klebsiella pneumoniae, Burkholderia mallei, Shigella boydii, Yersinia enterocolitica, Vibrio fischeri, and Pseudomonas putida[6] (Fig. 1a)
BcsQ has been shown to be essential for cellulose biogenesis in vivo and to localize at the polar site for cellulose secretion and cell-to-cell adhesion[11], BcsE is known to be necessary for maximal cellulose production[12] and we show here, by introducing non-polar deletions, that wild-type levels of BcsG and BcsR are essential for cellulose secretion (Supplementary Fig. 1b, c)
Summary
Secreted exopolysaccharides present important determinants for bacterial biofilm formation, survival, and virulence. Cellulose secretion typically requires the concerted action of a c-diGMP-responsive inner membrane synthase (BcsA), an accessory membrane-anchored protein (BcsB), and several additional Bcs components. Heterologous reconstitution and mutational analyses point toward a structure–function model, where accessory proteins regulate secretion by affecting both the assembly and stability of the system. These results lay the foundation for more comprehensive models of synthase-dependent exopolysaccharide secretion in biofilms and add a sophisticated secretory nanomachine to the diverse bacterial arsenal for virulence and adaptation. On the basis of these data, we propose functional models for nanoarray-like secretion of cellulose microfibers that would provide increased strength and biofilm forming capacity
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