Fig. 2: Surface remodeling underlies the spontaneous depletion-aggregation in VPS-producing culture.

a–c Phase diagrams were generated by mixing pVPS and VPS-producing cells (ΔABC cells) chemically fixed using 4% paraformaldehyde at (a) growth phase I (8 h), (b) growth phase II (20 h), and (c) growth phase I followed by incubation overnight with RbmB (0.05 mg/mL), at the indicated concentrations and visually scoring the cultures as either dispersed (filled circles) or aggregated (open circles). Red solid lines correspond to the phase boundary and dashed lines correspond to the original phase boundary obtained with 5Δ cells. Shown as an inset in each phase diagram is a schematic of the cell configuration for each cell type, respectively, created using BioRender.com. d, e Controllable transition of depletion-aggregation (images shown at z = 4 µm above the glass surface). Representative images were shown from four repeated experiments. d Cross-sectional confocal images of a strain carrying a plasmid with IPTG inducible expression of VC1086, a c-di-GMP phosphodiesterase (top) and an isogenic strain carrying a control plasmid with an inactive version of VC1086 (bottom). Both strains were grown in the presence of 1 µM IPTG overnight and visualized. Scale bar = 10 µm. e The strain carrying active VC1086 was grown for 15 h without IPTG (top) or without IPTG for 13 h followed by IPTG treatment for 2 h. Cell membranes were stained with FM 4-64. Scale bar = 10 µm. f Coarse-grained simulation of the depletion-aggregation phenomenon controlled by surface remodeling. Top: Snapshots of cellular arrangements following 100 million simulation steps, at varying VPS surface coverages (from left to right, 0%, 50%, and 100%). Individual VPS polymers are shown as green spheres. Patches on bacterial cell surfaces are colored depending on whether they are covered with VPS (green) or not (red). Bottom: The corresponding radial distribution function (RDF). Arrow indicates an emerging peak characteristic of depletion-induced parallel arrangement of the rod-shaped cells. g Simulated phase diagrams as a function of VPS surface coverage scored as either dispersed (filled circles) or aggregated (open circles). The ranges considered for the number of cells, number of polymer beads (pb), and VPS surface coverage are from 6 to 45 cells, 30,000 to 100,000 beads, and 0 to 100% VPS surface coverage, respectively, in a simulation box of 6.2 × 6.2 × 6.2 µm³. Solid color line from red (0% VPS coverage) to green (100% VPS surface coverage) in each phase diagram shows the boundary between the aggregated and dispersed phases. Source data are provided as a Source Data file.