Figures and data
A. Schematic diagram of the chemotaxis signal transduction network and flagellar motor of P. aeruginosa. Fusion protein CheY-EYFP and fluorescently labeled flagellar filaments were used as markers to indicate the position of the chemosensory array and motor, respectively. B. The labeling mechanism of flagellar filaments and chemotaxis regulatory protein CheY. Filaments (with cysteine point mutation FliCT394C) were labeled through sulfhydryl-maleimide conjugation, and cheY-eyfp fusion with a 3× glycine linker was used to visualize chemosensory array positions. C. Subcellular localization of CheY-EYFP in the wild-type strain of P. aeruginosa. CheY-EYFP is mainly located at the single cell pole, and the white arrow points to individuals with obvious chemosensory arrays at both cell poles, which generally have a large aspect ratio of the cell body. The yellow dashed box marks the cell outline. D. The merged imaging of flagellar filaments and CheY-EYFP in the wild-type strain of P. aeruginosa, where flagellar motor and chemosensory array colocalize in cells. White arrows point to individuals about to be divided, and the yellow dashed box marks the cell outline. The scale bar is 1 μm.
A. Subcellular localization of CheY-EYFP in the ΔflhF strain of P. aeruginosa. CheY-EYFP is no longer robustly distributed at the single cell pole. The yellow dashed box marks the cell outline. B. The merged imaging of flagellar filaments and CheY-EYFP in the ΔflhF strain of P. aeruginosa, flagellar motor and chemosensory array still colocalize in cells. The yellow dashed box marks the cell outline. The scale bar is 1 μm.
A. Subcellular localization of CheY-EYFP in various P. aeruginosa strains. The scale bar is 10 μm. B. Western blot analysis was performed to detect CheY expression in various P. aeruginosa strains. β-actin was used as the reference protein. C. Occurrence probability of obvious chemosensory arrays in P. aeruginosa wild-type and several mutant strains. The proportion of individuals with obvious chemosensory arrays decreased significantly in the motor-incomplete strains (ΔfliF and ΔfliG), and this value was further reduced after flhF knockout (ΔfliFΔflhF). The ΔfliF and ΔmotAΔmotCD strains have a similar chemosensory array occurrence probability as the wild-type strain. D. Absolute fluorescence brightness of the chemosensory array in P. aeruginosa wild-type and several mutant strains.
A. The evolution of cell aggregation as the intracellular CheY concentration increases by induction with higher concentrations of arabinose. The scale bar is 10 μm. B. Quantitative characterization of intracellular c-di-GMP levels at different CheY concentrations. From top to bottom, they correspond to the ΔcheY strain (N=198), wildtype strain (N=249) and CheY overexpression strain (N=228), respectively.
A. The construction mode of the chemotaxis network and flagellar motor throughout the complete cell growth cycle. B. The proximal growth mode of the P. aeruginosa flagellar motor and receptor clusters will effectively regulate the spatial range of CheY action, thus avoiding unintentional cross-pathway regulation.
