Defining the Layers of a Sensory Cilium With Storm and Cryoelectron Nanoscopy

Authors

Michael A Robichaux
Valencia L Potter
Zhixian Zhang
Feng He
Jun Liu
Michael F Schmid
Theodore G Wensel

Publication Date

11-19-2019

Journal

Proceedings of the National Academy of Sciences of the United States of America

DOI

10.1073/pnas.1902003116

PMID

31690665

PMCID

PMC6876244

PubMedCentral® Posted Date

11-5-2019

PubMedCentral® Full Text Version

Post-print

Published Open-Access

no

Keywords

Animals, Axoneme, Bardet-Biedl Syndrome, Centrioles, Cilia, Cryoelectron Microscopy, Disease Models, Animal, Electron Microscope Tomography, Eye Proteins, Mice, Mice, Inbred C57BL, Microscopy, Fluorescence, Microtubule-Associated Proteins, Microtubules, Multiprotein Complexes, Muscle Proteins, Nanotechnology, Photoreceptor Connecting Cilium, Qa-SNARE Proteins, Rod Cell Outer Segment, Single Molecule Imaging, Tumor Suppressor Proteins, primary cilia, photoreceptors, ciliopathies, superresolution, cryoelectron tomography

Abstract

Primary cilia carry out numerous signaling and sensory functions, and defects in them, "ciliopathies," cause a range of symptoms, including blindness. Understanding of their nanometer-scale ciliary substructures and their disruptions in ciliopathies has been hindered by limitations of conventional microscopic techniques. We have combined cryoelectron tomography, enhanced by subtomogram averaging, with superresolution stochastic optical reconstruction microscopy (STORM) to define subdomains within the light-sensing rod sensory cilium of mouse retinas and reveal previously unknown substructures formed by resident proteins. Domains are demarcated by structural features such as the axoneme and its connections to the ciliary membrane, and are correlated with molecular markers of subcompartments, including the lumen and walls of the axoneme, the membrane glycocalyx, and the intervening cytoplasm. Within this framework, we report spatial distributions of key proteins in wild-type (WT) mice and the effects on them of genetic deficiencies in 3 models of Bardet-Biedl syndrome.

Comments

Associated Data