Figure 5. Analysis of motility defects in PF6 Δaa transformants in the absence of the outer dynein arms.

(A) Western blots of axonemes (10 μg per lane) isolated from transformed pf6pf28 strains, probed with antibodies generated against PF6, IC69, and IC138. IC69 is an outer arm dynein intermediate chain. IC138 is an inner dynein arm intermediate chain. (B) Beat frequencies are significantly rescued in pf6pf28 strains expressing PF6ΔN or PF6ΔM (**p < 0.001, Student’s t test). (C) Quantification of stroke timing indicates that the recovery stroke is significantly longer in PF6ΔN and PF6ΔM mutants (**p < 0.001, Student’s t test). (D) Mean microtubule sliding velocities are not significantly different in pf6pf28 strains expressing PF6ΔN or PF6ΔM. (E) Waveform tracings indicate that waveform shape is similar in all mutants. The beginning of the effective stoke was designated at the point when the proximal end of the flagellum first began to reverse direction moving downward. By this criterion the effective stroke is initiated before the recovery stroke has reached the end of the flagellum. The same criterion was used to establish the beginning of the recovery stroke – the point at which the proximal portion of the flagellum begins to reverse direction. As is true for the effective stroke, the recovery stroke begins before the tip of the flagellum has completed the effective stroke. Each individual flagellar trace represents one frame of video separated by 2 ms. Red tracings indicate the recovery stroke. N ≥ 50 for all beat frequency and sliding velocity measurements. N ≥ 40 strokes and 10 cells for stroke time measurements. Total N is derived from three independent experiments. Error bars indicate SEM. Note that because the recovery stroke is delayed for some mutants, red tracings may overlap and, therefore, appear less numerous.