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. Author manuscript; available in PMC: 2021 Apr 6.
Published in final edited form as: Dev Cell. 2020 Feb 27;53(1):60–72.e4. doi: 10.1016/j.devcel.2020.01.029

Figure 2. Kinesin-3 access to the microtubule lattice is differentially gated by MAP7, tau, MAP2C, DCX, DCLK1, and MAP9.

Figure 2.

(A) TIRF-M images and kymographs of 1 nM KIF1A-mScarlet (kinesin-3) + 1 mM ATP in the absence and presence of 50 nM sfGFP-MAP7, 50 nM sfGFP-tau, 50 nM sfGFP-MAP2C, 100 nM DCX-sfGFP, 50 nM sfGFP-DCLK1, or 50 nM sfGFP-MAP9. Scale bars: 1 μm (x), 5 sec (y). (B) Quantification of the number of processive KIF1A-mScarlet motors + 1 mM ATP in the absence and presence of each MAP or MAP combination (means ± s.d. in motors μm−1min−1nM−1 are: 6.46 ± 3.09 for KIF1A alone (n=54 kymographs from 3 independent trials), 0.26 ± 0.33 for KIF1A + MAP7 (n=75 kymographs from 3 independent trials), 0.36 ± 0.64 for KIF1A + tau (n=71 kymographs from 3 independent trials), 0.13 ± 0.21 for KIF1A + MAP2C (n=69 kymographs from 3 independent trials), 3.77 ± 1.36 for KIF1A + DCX (n=65 kymographs from 3 independent trials), 4.61 ± 1.97 for KIF1A + DCLK1 (n=83 kymographs from 3 independent trials), 8.10 ± 2.61 for KIF1A + MAP9 (n=55 kymographs from 3 independent trials), 0.19 ± 0.26 for KIF1A + MAP7 + DCX (n=66 kymographs from 2 independent trials), 0.30 ± 0.25 for KIF1A + MAP7 + DCLK1 (n=46 kymographs from 2 independent trials), 0.33 ± 0.40 for KIF1A + MAP7 + MAP9 (n=76 kymographs from 2 independent trials), and 0.006 ± 0.004 for 50x the concentration of KIF1A (50 μM) + MAP2C (n=19 kymographs from 2 independent trials). All datapoints are plotted with lines indicating means ± s.d. P < 0.0001 (***) and P = 0.0034 (**) calculated by one-way ANOVA with Bonferroni correction. (C) Velocity histograms of KIF1A + 1 mM ATP in the absence and presence of DCX, DCLK1, and MAP9 with Gaussian fits. Means ± s.d. are 1252.5 ± 279.7, 931.0 ± 177.0, 1070.6 ± 280.6, and 1023.8 ± 293.6 nm/sec for KIF1A alone, KIF1A + DCX, KIF1A + DCLK1, and KIF1A + MAP9, respectively. P < 0.0001 for KIF1A vs. KIF1A + each MAP calculated by one-way ANOVA with Bonferroni correction. n=179, 77, 80 and 86 KIF1A motors for KIF1A alone, KIF1A + DCX, KIF1A + DCLK1, and KIF1A + MAP9, respectively from 2 independent trials each. (D) TIRF-M images and kymographs of 1 nM KIF1A-mScarlet + 1 mM ATP in the presence of 50 nM BFP-MAP7 (pink) with 50 nM DCX-sfGFP (green), or 50 nM BFP-MAP7 (pink) with 50 nM sfGFP-MAP9 (green). Scale bars: 1 μm (x), 5 sec (y).