Fig. 1: PopZ phase separates in Caulobacter crescentus.

a PopZ self-assembles at the poles of wildtype Caulobacter cells. A fluorescent image of ΔpopZ Caulobacter cells expressing mCherry-PopZ (red) from the xylX promoter on a high copy plasmid overlaid on a corresponding phase-contrast image. Scale bar, 1 μm. b The PopZ microdomain excludes ribosomes and forms a sharp convex boundary. (left) Slice through a tomogram of a cryo-ET focused ion beam-thinned ΔpopZ Caulobacter cell overexpressing mCherry-PopZ. A dashed red line shows the boundaries of the PopZ region. (right) Segmentation of the tomogram in (left) showing the outer membrane (dark brown), inner membrane (light brown), and ribosomes (gold). Scale bar, 1 μm. c, d PopZ creates droplets in deformed Caulobacter cells. c A fluorescent image of Caulobacter cells bearing a mreB A325P mutant, expressing mCherry-PopZ (red) from the xylX promoter on a high copy plasmid overlaid on a corresponding phase-contrast image. Scale bar, 1 μm. d Fluorescent images show the PopZ microdomain (red) extending into the cell body, concurrent with the thinning of the polar region, producing a droplet that dynamically moves throughout the cell. Frames are 2 min apart. Scale bar, 1 μm. e PopZ dynamics are not affected by a release from the cell pole. Recovery following targeted photobleaching of a portion of an extended PopZ microdomain in wildtype and mreB A325P mutant cells. Cells expressing mCherry-PopZ from a high copy plasmid were imaged for 12 frames of laser scanning confocal microscopy following targeted photobleaching with high-intensity 561 nm laser light. Shown is the mean ± SEM of the normalized fraction of recovered signal in the bleached region; n equals 15 cells.