Fig. 3. TbLC1 knockdown causes reverse cell motility. (A) Time-lapse series taken from video clips of induced (+Tet, 26 hpi) or uninduced (–Tet) TbLC1 knockdown cells. The dashed line marks the cell posterior (P) at the start of the time-lapse series. The black arrows in the first panel represent the direction expected for wild-type movement, with the anterior end (A) leading. Red arrows at the bottom show the actual direction of cellular movement. TbLC1 mutants move backward, with the posterior end leading. Significant cytokinesis problems prevented detailed motility analysis at later time points, but single cells observed at 49 hpi continued to exhibit reverse beat and reverse motility (not shown). Bars, 10 µm. (B) Cell traces (Baron et al., 2007) of uninduced (–Tet) and induced (+Tet, 26 hpi) TbLC1 knockdown cells. Lines show the distance traveled by cells over a 30-second interval. n=47 (–Tet) or 52 (+Tet) cells. Bars, 50 µm. Cell classifications are described in Materials and Methods. Inset pie charts display the percentage of each cell type observed in each strain. (C) Time-lapse series shows beat direction in two flagellar mutants that are stuck to slides. TbLC1 mutants (24 hpi) display a flagellar beat that originates at the base of the flagellum and is propagated to the tip (supplementary material Movie 3). Trypanin mutants (5 days post infection) display a tumbling phenotype (Hutchings et al., 2002) and maintain the wild-type tip-to-base flagellar waveform movement. Black arrows show the waveform position at the beginning of the series. White arrows show the position of the waveform at each time point.