Long and short range heterogeneity of amyloid curli and cellulose production is essential for morphogenesis and structural stability of Escherichia coli macrocolony biofilms

SERRA, DIEGO O.; YOUSEF, KAVEH; VON KLEIST, MAX; HENGGE, REGINE

Chicago

Conferencia; 7th ASM Conference on Biofilms; 2015

ASM Conferences; Fitnat Yildiz & Jean Marc Ghigo (Chairs).

In E. coli macrocolonies the emergence of elaborated structures, such as ridges and wrinkles, relies on a precise spatial distribution of flagella, amyloid curli fibers and cellulose that confer an optimal balance of cohesiveness and elasticity. Over a long range, this spatial matrix distribution is based on E. coli differentiating into two fundamentally different physiological layers in response to nutrient gradients building up in the biofilm: vegetatively growing cells entangled by flagella in the bottom layer (i.e., close to the nutrient-providing agar) and starving stationary-phase cells producing cellulose and curli fibers in the top layer. By combining macrocolony cryo-sectioning, specific labelling of matrix components and fluorescence and SEM microscopy, we found that the lower zone of the stationary-phase layer represents a highly heterogeneous transition area with cells that have switched to stationary phase and the production of mainly cellulose being located immediately adjacent to naked, elongated and flagellated cells. By contrast, further up in the stationary-phase layer, cellulose and curli form a dense nanocomposite that surrounds essentially all cells. Consistent with this, the expression of csgD, which encodes the biofilm regulator essential for curli and cellulose synthesis, is heterogeneous in the lower stationary-phase zone and relatively homogeneous in the contiguous upper zone. Importantly, we found that PdeR (YciR), a trigger enzyme with c-di-GMP degrading phosphodiesterase activity which acts as a central switching device in the control of csgD expression, is a key player in generating this short range matrix heterogeneity in the lower zone of the stationary phase layer. A knockout mutation in pdeR resulted in confluent and homogeneous distribution of cellulose and curli in the entire stationary phase layer. Mathematical modelling verified the potential of this PdeR-dependent switch to confer CsgD/curli/cellulose heterogeneity manifested as bistable states. Strikingly, as a consequence of losing the ability to produce matrix heterogeneously the pdeR mutant developed into large, rigid and extremely flat macrocolonies that, instead of buckling up into ridge or wrinkles, show scarred breaks at the macrocolony surface, thus reflecting extreme cohesiveness but reduced elasticity of the macrocolony. Overall, these results highlight that heterogeneity in matrix synthesis is essential for the development of complex morphological patterns in macrocolony biofilms.