Cryo-EM of dynein microtubule-binding domains shows how an axonemal dynein distorts the microtubule
- MRC Laboratory of Molecular Biology, United Kingdom
Abstract
Dyneins are motor proteins responsible for transport in the cytoplasm and the beating of axonemes in cilia and flagella. They bind and release microtubules via a compact microtubule-binding domain (MTBD) at the end of a coiled-coil stalk. We address how cytoplasmic and axonemal dynein MTBDs bind microtubules at near atomic resolution. We decorated microtubules with MTBDs of cytoplasmic dynein-1 and axonemal dynein DNAH7 and determined their cryo-EM structures using helical Relion. The majority of the MTBD is rigid upon binding, with the transition to the high affinity state controlled by the movement of a single helix at the MTBD interface. DNAH7 contains an 18-residue insertion, found in many axonemal dyneins, that contacts the adjacent protofilament. Unexpectedly we observe that DNAH7, but not dynein-1, induces large distortions in the microtubule cross-sectional curvature. This raises the possibility that dynein coordination in axonemes is mediated via conformational changes in the microtubule.
Data availability
CryoEM maps have been deposited to the EMDB and are instructed to be released on publication under codes 10060 and 10061PDB models have been deposited to the PDB and are instructed to be released on publication under codes 6RZA and 6RZB
Article and author information
Author details
Funding
Wellcome (WT210711)
- Andrew P Carter
Medical Research Council (MC_UP_A025_1011)
- Andrew P Carter
Medical Research Council (MC_UP_A025_1011)
- Sjors HW Scheres
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Samara L Reck-Peterson, University of California, San Diego, United States
Version history
- Received: March 26, 2019
- Accepted: July 1, 2019
- Accepted Manuscript published: July 2, 2019 (version 1)
- Version of Record published: July 15, 2019 (version 2)
Copyright
© 2019, Lacey et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Cryo-EM of dynein microtubule-binding domains shows how an axonemal dynein distorts the microtubule
eLife 8:e47145.
https://doi.org/10.7554/eLife.47145
Further reading
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- Cell Biology
- Structural Biology and Molecular Biophysics
Acetylation of α-tubulin at the lysine 40 residue (αK40) by αTAT1/MEC-17 acetyltransferase modulates microtubule properties and occurs in most eukaryotic cells. Previous literatures suggest that acetylated microtubules are more stable and damage resistant. αK40 acetylation is the only known microtubule luminal post-translational modification site. The luminal location suggests that the modification tunes the lateral interaction of protofilaments inside the microtubule. In this study, we examined the effect of tubulin acetylation on the doublet microtubule (DMT) in the cilia of Tetrahymena thermophila using a combination of cryo-electron microscopy, molecular dynamics, and mass spectrometry. We found that αK40 acetylation exerts a small-scale effect on the DMT structure and stability by influencing the lateral rotational angle. In addition, comparative mass spectrometry revealed a link between αK40 acetylation and phosphorylation in cilia.
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The dimeric two-pore OSCA/TMEM63 family has recently been identified as mechanically activated ion channels. Previously, based on the unique features of the structure of OSCA1.2, we postulated the potential involvement of several structural elements in sensing membrane tension (Jojoa-Cruz et al., 2018). Interestingly, while OSCA1, 2, and 3 clades are activated by membrane stretch in cell-attached patches (i.e. they are stretch-activated channels), they differ in their ability to transduce membrane deformation induced by a blunt probe (poking). Here, in an effort to understand the domains contributing to mechanical signal transduction, we used cryo-electron microscopy to solve the structure of Arabidopsis thaliana (At) OSCA3.1, which, unlike AtOSCA1.2, only produced stretch- but not poke-activated currents in our initial characterization (Murthy et al., 2018). Mutagenesis and electrophysiological assessment of conserved and divergent putative mechanosensitive features of OSCA1.2 reveal a selective disruption of the macroscopic currents elicited by poking without considerable effects on stretch-activated currents (SAC). Our results support the involvement of the amphipathic helix and lipid-interacting residues in the membrane fenestration in the response to poking. Our findings position these two structural elements as potential sources of functional diversity within the family.
