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

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

doi:10.1016/j.cell.2021.10.007

Cited by 79 publications

(197 citation statements)

References 137 publications

(196 reference statements)

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“…As discussed, exciting recent data has also emerged from studies of cilia MTs that demonstrates the ability of cryo-EM to identify macromolecules de novo within in situ or ex vivo preparations (Gui et al, 2021;Kiesel et al, 2020;Wang et al, 2021). The ability to faithfully localise and identify macromolecules on MTs in situ is likely to be a prominent target in the future direction of the field.…”

Section: Discussionmentioning

confidence: 99%

“…Finally, alongside work on single MTs and their binding proteins, there has been a recent flurry of exciting cryo-ET/ STA (Jordan et al, 2018;Zabeo et al, 2018;Owa et al, 2019;Greenan et al, 2020) and single-particle cryo-EM (Ma et al, 2019;Khalifa et al, 2020;Gui et al, 2021;Rao et al, 2021;Walton et al, 2021) work on the axonemal MT doublet structures of primary and motile cilia (Figure 2B). These studies have utilised both intact in situ and reduced membranated or de-membranated ex vivo preparations and have revealed a wealth of information on gross cilia architecture and the organisation, identities, and structure of axonemal MIPs, dynein complexes and intraflagellar transport trains (IFTs).…”

Section: High-resolution Studies Of Mts Using Cryo-emmentioning

confidence: 99%

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Structure Determination of Microtubules and Pili: Past, Present, and Future Directions

Garnett

Atherton

2022

Front. Mol. Biosci.

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Historically proteins that form highly polymeric and filamentous assemblies have been notoriously difficult to study using high resolution structural techniques. This has been due to several factors that include structural heterogeneity, their large molecular mass, and available yields. However, over the past decade we are now seeing a major shift towards atomic resolution insight and the study of more complex heterogenous samples and in situ/ex vivo examination of multi-subunit complexes. Although supported by developments in solid state nuclear magnetic resonance spectroscopy (ssNMR) and computational approaches, this has primarily been due to advances in cryogenic electron microscopy (cryo-EM). The study of eukaryotic microtubules and bacterial pili are good examples, and in this review, we will give an overview of the technical innovations that have enabled this transition and highlight the advancements that have been made for these two systems. Looking to the future we will also describe systems that remain difficult to study and where further technical breakthroughs are required.

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Section: Discussionmentioning

confidence: 99%

Section: High-resolution Studies Of Mts Using Cryo-emmentioning

confidence: 99%

Structure Determination of Microtubules and Pili: Past, Present, and Future Directions

Garnett

Atherton

2022

Front. Mol. Biosci.

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“…This ribbon diagram was generated with the PyMol molecular graphics system (Schrödinger) using Protein Data Bank accession no. 7RRO ( Gui et al, 2021 ).…”

Section: Axonemal Dyneinsmentioning

confidence: 99%

“…The ODA-DC in C. reinhardtii consists of three protein subunits, encoded by DCC1 (DC1), DCC2 (DC2), and DLE3 (DC3). In mammals, it consists of five protein subunits ( Gui et al, 2021 ) encoded by five genes, now named ODAD1 , ODAD2 , ODAD3 , ODAD4 , and CLXN (calaxin; Fig. 2 a and Fig.…”

Section: Axonemal Dyneinsmentioning

confidence: 99%

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Consensus nomenclature for dyneins and associated assembly factors

Braschi

Omran

Witman

et al. 2022

Journal of Cell Biology

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Dyneins are highly complex, multicomponent, microtubule-based molecular motors. These enzymes are responsible for numerous motile behaviors in cytoplasm, mediate retrograde intraflagellar transport (IFT), and power ciliary and flagellar motility. Variants in multiple genes encoding dyneins, outer dynein arm (ODA) docking complex subunits, and cytoplasmic factors involved in axonemal dynein preassembly (DNAAFs) are associated with human ciliopathies and are of clinical interest. Therefore, clear communication within this field is particularly important. Standardizing gene nomenclature, and basing it on orthology where possible, facilitates discussion and genetic comparison across species. Here, we discuss how the human gene nomenclature for dyneins, ODA docking complex subunits, and DNAAFs has been updated to be more functionally informative and consistent with that of the unicellular green alga Chlamydomonas reinhardtii, a key model organism for studying dyneins and ciliary function. We also detail additional nomenclature updates for vertebrate-specific genes that encode dynein chains and other proteins involved in dynein complex assembly.

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Ciliary dynein arms: Cytoplasmic preassembly, intraflagellar transport, and axonemal docking

Qiu

Roy

2022

Journal Cellular Physiology

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The microtubular scaffold of motile cilia-the axoneme, is decorated with dynein arms, which are large multiprotein complexes essential for ciliary motility. Dynein arms are arranged along the length of the axoneme in a precise repeating pattern, converting chemical energy from ATP hydrolysis into ciliary mechanical movement.How these complicated molecular machines are assembled coordinately and accurately, starting from mere polypeptide chains in the cytoplasm, remains a fascinating yet perplexing question. Rapidly emerging evidence, from multiple studies carried out with different model organisms and with various methodologies, has highlighted the existence of a dedicated assembly pathway. Here, we summarize recent progress made in clarifying the axonemal dynein arm assembly process, focusing on individual assembly steps, including cytoplasmic preassembly, intraflagellar transport, and axonemal docking. K E Y W O R D S dynein arm assembly, dynein arms, motile cilia 1 | INTRODUCTION Motile cilia are microtubule-based organelles that appear as hair-like protrusions on the cell surface and are present in diverse organisms across many evolutionary lineages. The microtubule scaffold of the cilium, the axoneme, is typically organized in a unique "9 + 2" pattern, in which a pair of central microtubules is surrounded by nine peripheral microtubule doublets (each doublet comprising a complete A and an incomplete B tubule) in a circular arrangement (Figure 1) (Ishikawa et al., 2021; Ishikawa, 2017). Importantly, a variety of biological processes, ranging from the free movement of individual cells such as protozoans like Paramecium and spermatozoa to the flow of mucus over the surface of the respiratory tract and circulation of cerebrospinal fluid within the brain ventricular cavities of mammals, all depend on the coordinated beating of motile cilia (F. Zhou & Roy, 2015).Ciliary motility is imparted by molecular motors known as outer and inner dynein arms (ODAs and IDAs, respectively), which are stably attached to the surface of the A tubule of the peripheral axonemal microtubules (Figure 1) (Ishikawa et al., 2021; Ishikawa, 2017). Dynein arms are very large, complex macromolecular assemblies that consist of multiple protein subunits (the estimated size can range from 1-to 2-MDa), overall contributing for ATP catalysis, microtubule binding activity, as well as regulatory input (King, 2016). The core catalytic subunits are comprised of dynein heavy chains (HCs), which have a length of around 4500 amino acid residues (around 500 KDa in size). Additionally, HCs associate and complex with multiple intermediate chains (ICs) and light chains (LCs), forming the dynein motors. Moreover, with the help of docking complexes (DC), dynein motors are able to stably associate with the axonemal microtubules, thereby enabling sliding motion of adjacent microtubule doublets and producing the overall oscillatory movement of the axoneme.Due to their elaborate structure, absence or mutation of even a single structural component of the dynein...

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

Cited by 79 publications

References 137 publications

Structure Determination of Microtubules and Pili: Past, Present, and Future Directions

Structure Determination of Microtubules and Pili: Past, Present, and Future Directions

Consensus nomenclature for dyneins and associated assembly factors

Ciliary dynein arms: Cytoplasmic preassembly, intraflagellar transport, and axonemal docking

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