Dynactin p150 promotes processive motility of DDB complexes by minimizing diffusional behavior of dynein

ABSTRACT: Cytoplasmic dynein is activated by forming a complex with dynactin and the adaptor protein BicD2. We used interferometric scattering (iSCAT) microscopy to track dynein–dynactin–BicD2 (DDB) complexes in vitro and developed a regression-based algorithm to classify switching between processive, diffusive, and stuck motility states. We find that DDB spends 65% of its time undergoing processive stepping, 4% undergoing 1D diffusion, and the remaining time transiently stuck to the microtubule. Although the p150 subunit was previously shown to enable dynactin diffusion along microtubules, blocking p150 enhanced the proportion of time DDB diffused and reduced the time DDB processively walked. Thus, DDB diffusive behavior most likely results from dynein switching into an inactive (diffusive) state, rather than p150 tethering the complex to the microtubule. DDB–kinesin-1 complexes, formed using a DNA adapter, moved slowly and persistently, and blocking p150 led to a 70 nm/s plus-end shift in the average velocity of the complexes, in quantitative agreement with the shift of isolated DDB into the diffusive state. The data suggest a DDB activation model in which dynactin p150 enhances dynein processivity not solely by acting as a diffusive tether that maintains microtubule association, but rather by acting as an allosteric activator that promotes a conformation of dynein optimal for processive stepping. In bidirectional cargo transport driven by the opposing activities of kinesin and dynein–dynactin–DDB, the dynactin p150 subunit promotes retrograde transport and could serve as a target for regulators of transport.

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Work Title Dynactin p150 promotes processive motility of DDB complexes by minimizing diffusional behavior of dynein
Access
Open Access
Creators
  1. QINGZHOU FENG
  2. William O Hancock
Keyword
  1. DDB, p150, activation, algorithm, bidirectional transport
License Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0)
Work Type Dataset
Publication Date 2020
DOI doi:10.26207/8q5t-rf37
Deposited March 04, 2020

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Version 1
published

  • Created
  • Added D1203slidingP1.m
  • Added DDB_32.mat
  • Added landing.xlsx
  • Added controlAntibody.xlsx
  • Added README.txt
  • Added NoSingle1205P3.m
  • Added Apolock.xlsx
  • Added p150_32.mat
  • Added Dcompare.pzfx
  • Added middle1205_2.m
  • Added Creator QINGZHOU FENG
  • Added Creator William O Hancock
  • Published