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. 2017 Oct 2;216(10):3051–3060. doi: 10.1083/jcb.201703201

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© 2017 Huynh and Vale

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/).

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Figure 1. — BICD2_FL activation by Rab6a^GTP. (A) Domain architecture of mammalian Bicaudal D2. N188T, I189F, and S107L are mutations associated with human SMA-LED, and F739I is the analogous mutation to the classical Drosophila mutation F684I. Amino acids 400 and 594 indicate the end of constructs used in this study. (B) Porcine brain lysate pull-down using full-length (FL) BICD2 as bait. Increasing amounts of recombinant Rab6a^GTP (left) or Rab6a^GDP (right) were added to the lysate, and the amount of endogenous dynein (dynein intermediate chain [DIC]) and dynactin (p150 subunit) bound was analyzed by immunoblot. (C) Sample kymographs of single-molecule DDB (aa 25–400 or FL BICD2) motility on microtubules. Fluorescence is from the N-terminal sfGFP tag on BICD2. (D) Quantification of the number of moving motors per micrometer of microtubule; mean and SEM from n = 3 independent experiments (each experiment measuring a minimum of 30 microtubules). (E) Quantification of number of moving motors per micrometer of microtubule of BICD2_FL with either Rab6a^GDP or Rab6a^GTP added; mean and SEM from n = 3 independent experiments (each experiment measuring a minimum of 30 microtubules).