A c-di-GMP-dependent switch in the control of short-range heterogeneity of the biofilm regulator CsgD and extracellular matrix in Escherichia coli macrocolony biofilms

SERRA, DIEGO O.; HENGGE, REGINE

Schloss Hohenkammer

Conferencia; SPP1617 International Meeting 2019; 2019

Deutsche Forschungsgemeinschaft (DFG)

Morphogenesis of E. coli macrocolony biofilms relies on a precise spatial distribution of flagella, amyloid curli fibers and cellulose. Long-range heterogeneity of these matrix components is a consequence of stratification into different physiological layers in response to nutrient gradients: vegetatively growing, flagellated cells in the bottom layer (close to the nutrients) and starving stationary-phase cells producing cellulose and curli fibers in the top layer. Using microscopic approaches, we found that during maturation of a macrocolony a highly heterogeneous intermediate layer is formed where cells that have switched to stationary phase and produce mainly cellulose, are located immediately adjacent to matrix-free 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 intermediate zone and relatively homogeneous in the contiguous upper stationary phase zone. This spatial pattern in macrocolonies correlates with heterogeneity of csgD expression during transition into stationary phase in liquid cultures, with bacteria exhibiting three discrete states: CsgDOFF, CsgDON high and CsgDON very high. Importantly, we found that PdeR and DgcM, i.e. two interacting enzymes that break and make c-di-GMP which are involved in controlling csgD transcription, act as a switching device generating csgD heterogeneity - and hence heterogeneity of matrix production - in response to global and local c-di-GMP levels controlled mainly by PdeH/DcgE and DgcM, respectively. Strikingly, by losing the ability to produce matrix heterogeneously, pdeR and pdeR/dgcM mutants developed into large, rigid and extremely flat macrocolonies that show scarred breaks at the surface, thus reflecting extreme cohesiveness but reduced elasticity of the macrocolony. Overall, these results highlight that heterogeneity in matrix synthesis is essential for morphogenesis and structural integrity of macrocolony biofilms, which explains, why E. coli regulates this process so tightly.