The University of Southampton
University of Southampton Institutional Repository

De novo identification of mammalian ciliary motility proteins using cryo-EM

De novo identification of mammalian ciliary motility proteins using cryo-EM
De novo identification of mammalian ciliary motility proteins using cryo-EM

Dynein-decorated doublet microtubules (DMTs) are critical components of the oscillatory molecular machine of cilia, the axoneme, and have luminal surfaces patterned periodically by microtubule inner proteins (MIPs). Here we present an atomic model of the 48-nm repeat of a mammalian DMT, derived from a cryoelectron microscopy (cryo-EM) map of the complex isolated from bovine respiratory cilia. The structure uncovers principles of doublet microtubule organization and features specific to vertebrate cilia, including previously unknown MIPs, a luminal bundle of tektin filaments, and a pentameric dynein-docking complex. We identify a mechanism for bridging 48- to 24-nm periodicity across the microtubule wall and show that loss of the proteins involved causes defective ciliary motility and laterality abnormalities in zebrafish and mice. Our structure identifies candidate genes for diagnosis of ciliopathies and provides a framework to understand their functions in driving ciliary motility.

axonemal dyneins, ciliary motility, ciliopathies, cryo-EM, doublet microtubules, microtubule inner proteins
0092-8674
5791-5806.e19
Gui, Miao
f55ad726-717a-4ad0-ad9b-8f43a0a2badb
Hannah, Farley
55f0b7d2-de7a-4df5-9514-6afa897d07fa
Anujan, Priyanka
1ad18e28-42d9-464f-8c27-2c18741fa771
Anderson, Jacob R.
91e48f52-287f-4fdc-b48a-2be368ba85eb
Thompson, James
a0a1e940-d720-47de-81d7-ebcd48738239
Lucas, Jane
5cb3546c-87b2-4e59-af48-402076e25313
Bingle, Colin D
9a639961-c20a-4da0-9ce1-ddb2b9e81a69
Norris, Dominic P.
14b7f8e4-162c-4218-be30-8396224464d1
Roy, Sudipto
8a0fc161-c107-4642-9148-3f70b363e83c
Brown, Alan
90defe2f-30f9-4e22-aaaa-3a6b5ab3f921
Gui, Miao
f55ad726-717a-4ad0-ad9b-8f43a0a2badb
Hannah, Farley
55f0b7d2-de7a-4df5-9514-6afa897d07fa
Anujan, Priyanka
1ad18e28-42d9-464f-8c27-2c18741fa771
Anderson, Jacob R.
91e48f52-287f-4fdc-b48a-2be368ba85eb
Thompson, James
a0a1e940-d720-47de-81d7-ebcd48738239
Lucas, Jane
5cb3546c-87b2-4e59-af48-402076e25313
Bingle, Colin D
9a639961-c20a-4da0-9ce1-ddb2b9e81a69
Norris, Dominic P.
14b7f8e4-162c-4218-be30-8396224464d1
Roy, Sudipto
8a0fc161-c107-4642-9148-3f70b363e83c
Brown, Alan
90defe2f-30f9-4e22-aaaa-3a6b5ab3f921

Gui, Miao, Hannah, Farley, Anujan, Priyanka, Anderson, Jacob R., Thompson, James, Lucas, Jane, Bingle, Colin D, Norris, Dominic P., Roy, Sudipto and Brown, Alan (2021) De novo identification of mammalian ciliary motility proteins using cryo-EM. Cell, 184 (23), 5791-5806.e19. (doi:10.1016/j.cell.2021.10.007).

Record type: Article

Abstract

Dynein-decorated doublet microtubules (DMTs) are critical components of the oscillatory molecular machine of cilia, the axoneme, and have luminal surfaces patterned periodically by microtubule inner proteins (MIPs). Here we present an atomic model of the 48-nm repeat of a mammalian DMT, derived from a cryoelectron microscopy (cryo-EM) map of the complex isolated from bovine respiratory cilia. The structure uncovers principles of doublet microtubule organization and features specific to vertebrate cilia, including previously unknown MIPs, a luminal bundle of tektin filaments, and a pentameric dynein-docking complex. We identify a mechanism for bridging 48- to 24-nm periodicity across the microtubule wall and show that loss of the proteins involved causes defective ciliary motility and laterality abnormalities in zebrafish and mice. Our structure identifies candidate genes for diagnosis of ciliopathies and provides a framework to understand their functions in driving ciliary motility.

Text
Gui_et_al_FINAL - Accepted Manuscript
Available under License Creative Commons Attribution.
Download (225kB)
Text
1-s2.0-S0092867421011818-main - Version of Record
Available under License Creative Commons Attribution.
Download (11MB)

More information

Accepted/In Press date: 7 October 2021
Published date: 11 November 2021
Additional Information: Funding Information: Cryo-EM data were collected at The Harvard Cryo-EM Center for Structural Biology. We thank Sarah Sterling, Richard Walsh, and Shaun Rawson for microscopy assistance; Ross Tomaino (Harvard Medical School) for mass spectrometry analysis; Y.L. Chong (Institute of Molecular and Cell Biology) for identification of zebrafish mutants; the A ∗ STAR Microscopy Platform and the National University of Singapore EM unit for TEM analysis of zebrafish KV; staff at the Mary Lyon Centre for mouse husbandry, genotyping, and histology; Henrik Westerberg and Jenni Vibert (MRC Harwell Institute) for help with particle image velocimetry; Claire Jackson, Janice Coles, Liz Adams, and Peter Lackie (PCD Centre, University of Southampton) for assessing mouse tracheal ciliary beating and TEM images; Elspeth Bruford (HGNC) for gene nomenclature discussions; and Marie Bao for comments. P.A. and D.W.M. were supported by University of Sheffield-A ∗ STAR Singapore and University of Manchester-A ∗ STAR Singapore doctoral studentships, respectively. This work was supported by awards from the UK Medical Research Council ( MC_U142670370 to D.P.N.), A ∗ STAR Singapore ( SC15-R0010 to S.R.), National Institutes of Health ( NIGMS 1R01GM141109 ), the LouLou Foundation , the E. Matilda Ziegler Foundation for the Blind , the Smith Family Foundation , and the Pew Charitable Trusts (to A.B.). The National PCD Centre in Southampton is commissioned and funded by NHS England ; PCD research is supported by the National Institute for Health Research ( RfPB PB-PG-1215-20014 and 200470 ) and The AAIR Charity (Reg. No. 1129698 ). Funding Information: Cryo-EM data were collected at The Harvard Cryo-EM Center for Structural Biology. We thank Sarah Sterling, Richard Walsh, and Shaun Rawson for microscopy assistance; Ross Tomaino (Harvard Medical School) for mass spectrometry analysis; Y.L. Chong (Institute of Molecular and Cell Biology) for identification of zebrafish mutants; the A?STAR Microscopy Platform and the National University of Singapore EM unit for TEM analysis of zebrafish KV; staff at the Mary Lyon Centre for mouse husbandry, genotyping, and histology; Henrik Westerberg and Jenni Vibert (MRC Harwell Institute) for help with particle image velocimetry; Claire Jackson, Janice Coles, Liz Adams, and Peter Lackie (PCD Centre, University of Southampton) for assessing mouse tracheal ciliary beating and TEM images; Elspeth Bruford (HGNC) for gene nomenclature discussions; and Marie Bao for comments. P.A. and D.W.M. were supported by University of Sheffield-A?STAR Singapore and University of Manchester-A?STAR Singapore doctoral studentships, respectively. This work was supported by awards from the UK Medical Research Council (MC_U142670370 to D.P.N.), A?STAR Singapore (SC15-R0010 to S.R.), National Institutes of Health (NIGMS 1R01GM141109), the LouLou Foundation, the E. Matilda Ziegler Foundation for the Blind, the Smith Family Foundation, and the Pew Charitable Trusts (to A.B.). The National PCD Centre in Southampton is commissioned and funded by NHS England; PCD research is supported by the National Institute for Health Research (RfPB PB-PG-1215-20014 and 200470) and The AAIR Charity (Reg. No. 1129698). M.G. and J.J.B. purified samples, collected cryo-EM data, and picked particles with help from S.M. S.K.S. and Q.Z. M.G. processed the cryo-EM data. M.G. and J.R.A. built models and analyzed structures. H.F. and J.B.W. analyzed mouse situs and nodal and tracheal cilia motility. P.A. and D.W.M. created and characterized zebrafish mutants. T.Q. performed TEM of KV cilia. J.T. videoed and analyzed cilia motility and performed TEM of mouse tracheal cilia. J.S.L. C.D.B. D.P.N. S.R. and A.B. supervised the research. M.G. J.R.A. J.B.W. S.R. and A.B. wrote the manuscript with input from all authors. The authors declare no competing interests. Publisher Copyright: © 2021 The Author(s) Copyright: Copyright 2021 Elsevier B.V., All rights reserved.
Keywords: axonemal dyneins, ciliary motility, ciliopathies, cryo-EM, doublet microtubules, microtubule inner proteins

Identifiers

Local EPrints ID: 452211
URI: http://eprints.soton.ac.uk/id/eprint/452211
ISSN: 0092-8674
PURE UUID: 9a94c60a-3b66-4795-a406-2b6ab5c84db0
ORCID for James Thompson: ORCID iD orcid.org/0000-0002-9285-1317
ORCID for Jane Lucas: ORCID iD orcid.org/0000-0001-8701-9975

Catalogue record

Date deposited: 30 Nov 2021 17:31
Last modified: 06 Jun 2024 01:51

Export record

Altmetrics

Contributors

Author: Miao Gui
Author: Farley Hannah
Author: Priyanka Anujan
Author: Jacob R. Anderson
Author: James Thompson ORCID iD
Author: Jane Lucas ORCID iD
Author: Colin D Bingle
Author: Dominic P. Norris
Author: Sudipto Roy
Author: Alan Brown

Download statistics

Downloads from ePrints over the past year. Other digital versions may also be available to download e.g. from the publisher's website.

View more statistics

Atom RSS 1.0 RSS 2.0

Contact ePrints Soton: eprints@soton.ac.uk

ePrints Soton supports OAI 2.0 with a base URL of http://eprints.soton.ac.uk/cgi/oai2

This repository has been built using EPrints software, developed at the University of Southampton, but available to everyone to use.

We use cookies to ensure that we give you the best experience on our website. If you continue without changing your settings, we will assume that you are happy to receive cookies on the University of Southampton website.

×