Figure 7.
Cargo Transport and Force Generation by Growing and Shrinking MT Tips
(A) Schematics of the experimental setup.
(B) Kymographs showing MTs polymerized from HiLyte-488 tubulin, bio-mCherry-MTLS, and mCherry-EB3 and Qdot705-streptavidin. Notation above the kymographs describes the experimental conditions. See also Video S7.
(C) Setup of the optical trap experiments.
(D) An example of a bead trace in the presence of EB3 showing the raw signal at 1 kHz (gray) and the signal after smoothing with a running average of 100 points (red). Images at the bottom show corresponding frames from the DIC video (Video S8). The red cross marks the center of the optical trap.
(E) Analogous experiment in the absence of EB3. Bottom trace: the fast back and forth movements continued after the MT tip grew past the position of the trapped bead (white arrowhead).
(F) Example trace and image of a free, unattached bead in a trap.
(G and I) Force amplitude and force duration measured at the growing (G) or shrinking (I) MT tip, in the presence (black, n = 24 [G] or 19 [I]) or absence (red, n = 10 [G] and 22 [I]) of 100 nM EB3. Horizontal line: median; box: 25%–75%, whiskers: minimal and maximal values.
(H) Example trace of a Bio-mCherry-MTLS-coated bead in the absence of EB3 pulled by a shortening MT tip depicted as in (D).