Fig. 3: Distinct PBP2 localization in A. excentricus and C. crescentus and its correlation with active PG synthesis.

a Subcellular localization of mCherry-PBP2 in A. excentricus. Top: Representative images are shown (n  =  3 biological replicates). Left to right: Phase, mCherry-PBP2 fluorescence, and merged images. Scale bar: 2 µm. A heatmap of mCherry-PBP2 foci at the population level is displayed. In the heatmap, cells were oriented using the old pole labeled with WGA (cyan), with the white line indicating the midcell. Bottom: A demograph showing the localization of mCherry-PBP2 fluorescence at the population level, with cells arranged by length and oriented with the old pole towards the right. b Short-pulse FDAA (BADA) labeling of the A. excentricus mCherry-PBP2 strain. Top: A schematic depicting the experiment. Cells were labeled with 250 μM BADA for 5% of their generation time, fixed with 70% (v/v) ethanol, and imaged. Middle: Representative phase, fluorescence (mCherry-PBP2 and BADA), and merged images are shown (n  =  3 biological replicates). Scale bar: 2 µm. Bottom: Population-level demographs showing the localization of the fluorescence intensities of mCherry-PBP2, BADA, and their overlays. See Supplementary Fig. 4a for heatmaps and density maps of mCherry-PBP2 and BADA at the population level. c Short-pulse FDAA (TADA) labeling of the C. crescentus GFP-PBP2 strain. Top: A schematic depicting the experiment. Cells were labeled with 250 μM TADA for 5% of their generation time, fixed with 70% (v/v) ethanol, and imaged. Middle: Representative phase, fluorescence (GFP-PBP2 and TADA), and merged images are shown (n  =  3 biological replicates). Scale bar: 2 µm. Bottom: Population-level demographs showing the localization of the fluorescence intensities of GFP-PBP2, TADA, and their overlays. See Supplementary Fig. 4b, c for additional fluorescence images and population-level demographs showing the differences in localization of PBP2 in A. excentricus vs. C. crescentus.