Fig. 3. Effect of LIS1 and NudE on Dynein Motility.
(A) Histogram of single dynein travel distances alone (D), in the presence of LIS1 (molar ratio of D:L = 1:2), and in the presence of both NudE and LIS1 (D:N:L = 1:9:10). Exponential decay fits are shown in red, and decay constant ± SEM is indicated in each case. A small percentage of beads had bidirectional motion of 2 µm or more in each direction, did not spontaneously detach from microtubules, and were excluded from analysis; such beads showed no force production capacity in either direction and were deemed to be diffusive rather than processive. D and DL processivity values are similar, whereas DNL processivity is clearly increased (P <0.03). (B) Microtubule-binding activity for D, DN, and DNL beads. We quantify the percentage of beads with visible binding and travel events, held close to a microtubule in a weak optical trap (See Methods). NudE (molar ratio 1:9 D:N) potently reduced microtubule binding activity, an effect rescued by addition of LIS1 (1:9:2 D:N:L). Neither LIS1 nor NudE produced a comparable inhibition of binding activity in similar experiments with kinesin motors (Fig. S2B). Exact CI error bars are reported (see Supplement). (C) Individual traces of motor-driven movements along microtubules (D:L ratios indicated; D:N:L is as in A). D (black) and DNL (green) beads show robust motility whereas DL bead movements (dark and light red) are interrupted by pauses that result in a net slow-down of transport. The effect becomes more prominent as the LIS1 concentration increases. See Fig. S3 for more extended and extreme examples of bead travel. For all experiments, dynein was first adsorbed to beads, followed as needed by NudE and then by LIS1.