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. 2017 Nov 1;6:e28842. doi: 10.7554/eLife.28842

Figure 5. CleA tunes the flagellar motor by interfering with the chemotaxis response.

(a) Cle proteins impede spreading on semisolid agar. Relative spreading areas of C. crescentus wild type and fliM, cheY and cle mutants are indicated. Strains were incubated at 30°C and motility was scored after 72 hr as the overall area covered by individual strains. Strains containing single or multiple deletions are indicated. (b) Mutants lacking CleA show a hyper-reversal phenotype. Directional reversal frequencies and mean run speed were measured for individual swimming cells of C. crescentus wild type and mutants lacking CheB, CheR, CleA, or all five Cle proteins. All strains analyzed harbored an additional deletion in the pilA gene to avoid pili-mediated motility variations. Cells were located in a pseudo-2D environment and their swimming trajectories were recorded at 16 frames per second. The fraction of cells with a given reversal frequency (left) or mean run speed (right) is indicated. (c) CleA promotes smooth swimming by antagonizing the major C. crescentus CheY, CheYII. Analysis of reversal frequencies and mean speed of the strains indicated was as in (b). It should be noted that the reversal frequency profiles of strains ΔcheYII, DcheYIIΔcleA and ΔcheYIIΔcheYI overlay. Experiments in (b) and (c) include the analysis of >850 cells for each strain.

Figure 5.

Figure 5—figure supplement 1. Reversal frequencies and speed of C. crescentus mutants lacking specific CheY and Cle components.

Figure 5—figure supplement 1.

(a) Reversal frequencies (left) and mean run speeds (right) were determined as indicated in the legend to Figure 5b. Strains lacking CleD (ΔcleD) or CleB, CleC, CleD and CleE (ΔcleB-E) showed wild-type like reversal frequencies and speed. (b) Reversal frequencies and speed in cle mutants and mutants lacking all C. crescentus CheR methyltransferases or CheB methylesterases. Median (red bars) reversal frequencies and speeds are from trajectories of more than 2000 cells (WT = 6719 cells; ΔcleA = 4602 cells; ΔcleD = 3134 cells; ΔcleB-E = 2532 cells; ΔcleA-E = 3757 cells; ΔcheR = 5042 cells; ΔcheB = 2392 cells) combined from multiple independent experiments. Number of replicates is seven, five, four, two, four, three, and two, respectively. The edges of the box indicate 25th and 75th percentiles of the data set. (c) Functional interaction of CleA and CheYII. Reversal frequencies and speed were measured in the mutants indicated. Data are from more than 1000 cells (WT = 6719 cells; ΔcleA = 4602 cells; ΔcheYI = 2205 cells; ΔcheYII = 1347 cells; ΔcleAΔcheYII = 1762 cells; ΔcheYIΔcheYII = 1375 cells) combined from multiple independent experiments. Experiments were repeated twice except for WT and ΔcleA, which were repeated seven and five times, respectively. All strains harbored a deletion in pilA encoding the major pilin subunit.