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

Miao Gui,Hannah Farley,Priyanka Anujan,Jacob R Anderson,Dale W Maxwell,Jonathan B Whitchurch,J Josephine Botsch,Tao Qiu,Shimi Meleppattu,Sandeep K Singh,Qi Zhang,James Thompson,Jane S Lucas,Colin D Bingle,Dominic P Norris,Sudipto Roy,Alan Brown

doi:10.1016/j.cell.2021.10.007

Abstract

SUMMARYDynein-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.

Highlights

Motile cilia are eukaryotic organelles responsible for cellular locomotion and movement of extracellular fluids
A cryoelectron microscopy study of doublet microtubules (DMTs) from the biflagellate alga Chlamydomonas reinhardtii revealed that their luminal surfaces are patterned by a 48-nm repeating network of at least 33 different microtubule inner proteins (MIPs) (Ma et al, 2019)
A map of the 96-nm external repeat confirmed the overall periodicity of the MIP structure as 48 nm (Figure S1E), and comparison with the subtomogram average of bovine DMT (Greenan et al, 2020) showed that all prominent MIPs were retained (Figure S1F)

Summary

Introduction

Motile cilia are eukaryotic organelles responsible for cellular locomotion and movement of extracellular fluids. During vertebrate embryogenesis, motile cilia are responsible for the directional flow of extraembryonic fluids within the leftright organizer (LRO) that establishes left-right asymmetry of visceral organs like the heart (Nonaka et al, 1998). Motile cilia power the movement of spermatozoa and the flow of mucus in the respiratory system (reviewed in Zhou and Roy, 2015). Consistent with these varied functions, impairment of ciliary motility can cause laterality abnormalities, including congenital heart defects, infertility, and chronic respiratory disease, that often collectively manifest in the ciliopathy primary ciliary dyskinesia (PCD) (Legendre et al, 2021).

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