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Molecular architecture of the mammalian 2-oxoglutarate dehydrogenase complex
The 2-oxoglutarate dehydrogenase complex (OGDHc) is a rate-limiting enzyme complex regulating the TCA cycle. Here, the authors use cryo-EM and cryo-ET to discern native structures of the whole complex and reveal its assembly mechanism.
- Yitang Zhang
- , Maofei Chen
- & Maojun Yang
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Article
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Intracellular morphogenesis of diatom silica is guided by local variations in membrane curvature
The silica cell wall of unicellular algae has intricate architecture unattainable by current technology. In this work it is shown that membrane contact sites are the biological morphogenesis handles to shape such intracellular mineralization.
- Lior Aram
- , Diede de Haan
- & Assaf Gal
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Human cell surface-AAV interactomes identify LRP6 as blood-brain barrier transcytosis receptor and immune cytokine IL3 as AAV9 binder
Engineered adeno-associated viruses (AAVs) aim to improve safety and potency for use in gene therapy but mechanisms underlying these features are poorly understood. Here, authors use unbiased screens to identify an interaction with the human immune system and a determinant of enhanced brain potency.
- Timothy F. Shay
- , Seongmin Jang
- & Viviana Gradinaru
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In situ structural determination of cyanobacterial phycobilisome–PSII supercomplex by STAgSPA strategy
The authors solved the native structure of cyanobacterial PBS–PSII supercomplex at a resolution of 3.5 Å via STAgSPA strategy, revealing the association details and possible energy transfer pathways between PBS and PSII.
- Xing Zhang
- , Yanan Xiao
- & Sen-Fang Sui
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Cell-to-cell interactions revealed by cryo-tomography of a DPANN co-culture system
DPANN is a widespread and diverse group of archaeal obligate ectosymbionts that depend on their host for proliferation. Here, authors use cryo-ET and proteomics to reveal intercellular proteinaceous nanotubes to facilitate DPANN-host interactions.
- Matthew D. Johnson
- , Doulin C. Shepherd
- & Debnath Ghosal
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Helical superstructures between amyloid and collagen in cardiac fibrils from a patient with AL amyloidosis
Here the authors report the cryo-EM structure of heart-derived fibrils of an AL amyloidosis patient. Surprisingly, the fibrils form helical superstructures with collagen VI, potentially signifying a distinct pathophysiological mechanism for amyloidoses.
- Tim Schulte
- , Antonio Chaves-Sanjuan
- & Stefano Ricagno
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CryoET reveals actin filaments within platelet microtubules
Using focused ion beam milling and cryoelectron tomography, Tsuji et al. found actin filaments, with both cofilin-bound and canonical morphologies, within the lumen of human platelet microtubules and reconstituted these structures in vitro.
- Chisato Tsuji
- , Marston Bradshaw
- & Mark P. Dodding
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Assembly of respiratory syncytial virus matrix protein lattice and its coordination with fusion glycoprotein trimers
Here the authors use cryo-electron tomography and sub-tomogram averaging to determine the molecular organization of RSV. The structures show that the matrix (M) protein packs as a helical-like lattice and the F glycoprotein is present as anti-parallel pairs of trimers and coordinates with the M lattice.
- Bryan S. Sibert
- , Joseph Y. Kim
- & Elizabeth R. Wright
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Bactericidal effect of tetracycline in E. coli strain ED1a may be associated with ribosome dysfunction
The human gut contains diverse bacterial strains that are beneficial/critical for health. Here, the authors compare the response of human gut and laboratory E. coli strains to the antibiotic tetracycline in molecular detail and find a severe dysfunction of protein synthesis only in the gut strain.
- Iskander Khusainov
- , Natalie Romanov
- & Martin Beck
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Angle between DNA linker and nucleosome core particle regulates array compaction revealed by individual-particle cryo-electron tomography
Here, using cryo-ET, the 3D structures of individual nucleosome particles were characterized to observe changes under varying ionic strengths and in the presence of protein H1, revealing key regulatory roles in chromatin organization dynamics.
- Meng Zhang
- , César Díaz-Celis
- & Gang Ren
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High-confidence 3D template matching for cryo-electron tomography
Enabling visual proteomics with high-confidence 3D template matching (TM) in CryoET. 3D TM precisely localizes macromolecular complexes, individual subunits, and different functional states in situ, revealing molecular interactions within cells.
- Sergio Cruz-León
- , Tomáš Majtner
- & Gerhard Hummer
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Targeted small molecule inhibitors blocking the cytolytic effects of pneumolysin and homologous toxins
The pore-forming toxin pneumolysin is responsible for the high mortality seen in pneumococcal infections unresponsive to antibiotics. In this work, authors report a small molecule inhibitor targeting pneumolysin and related ones as an anti-virulence strategy protecting human cells during infection.
- Umer Bin Abdul Aziz
- , Ali Saoud
- & Jörg Rademann
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Ultrastructure of human brain tissue vitrified from autopsy revealed by cryo-ET with cryo-plasma FIB milling
Here the authors report a method for cryogenic electron microscopy imaging of human brain tissue samples directly obtained from autopsy, offering insights into cellular ultrastructure and a tool to study potential pathologic features.
- Benjamin C. Creekmore
- , Kathryn Kixmoeller
- & Yi-Wei Chang
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DeepETPicker: Fast and accurate 3D particle picking for cryo-electron tomography using weakly supervised deep learning
Picking particles of biological macromolecules is critical for solving their structures in situ using cryo-electron tomograms. Here, authors develop DeepETPicker, a deep learning-based tool for fast, accurate, and automated picking of three-dimensional particles.
- Guole Liu
- , Tongxin Niu
- & Ge Yang
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In situ structure of actin remodeling during glucose-stimulated insulin secretion using cryo-electron tomography
Actin mediates insulin secretion in pancreatic β-cells through remodeling. Here, authors report the in situ structure of actin remodeling and quantify changes in architecture, alignment, and interactions during glucose-stimulated insulin secretion.
- Weimin Li
- , Angdi Li
- & Liping Sun
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Local structural preferences in shaping tau amyloid polymorphism
In this work, using a combination of Cryo-EM, in-cell experiments and biophysical analysis, the authors decoded the aggregation propensity of tau, revealing 5 central hot spots in its primary sequence and identify PAM4 as short segment that determines both the structure, as well as the cellular propagation of tau aggregates extracted from Alzheimer’s disease, corticobasal degeneration, and progressive supranuclear palsy patients.
- Nikolaos Louros
- , Martin Wilkinson
- & Joost Schymkowitz
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Overcoming resolution attenuation during tilted cryo-EM data collection
Here, the authors quantify the effect of cryo-EM data acquisition with stage-tilt on the global resolution of reconstructions and present a tool for predicting an optimal stage-tilt angle to ameliorate the effects of preferred specimen orientation.
- Sriram Aiyer
- , Philip R. Baldwin
- & Dmitry Lyumkis
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Multi-scale structures of the mammalian radial spoke and divergence of axonemal complexes in ependymal cilia
Radial spokes (RS) are crucial in coordinating ciliary motility. Here, authors use cryo-EM and cryo-ET to gain insight into mammalian RS divergence in ependymal cilia, RS assembly mechanism and the structure-function relationships of ciliary and flagellar axonemes.
- Xueming Meng
- , Cong Xu
- & Yao Cong
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Targeted mutagenesis of the herpesvirus fusogen central helix captures transition states
The early stages of viral fusogen conformational change required for enveloped viruses to infect cells is unclear. Here, authors capture the herpesvirus fusogen, glycoprotein B, in early transitional states by cryo-EM.
- Momei Zhou
- , Benjamin Vollmer
- & Stefan L. Oliver
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SARS-CoV-2 nsp3 and nsp4 are minimal constituents of a pore spanning replication organelle
Coronavirus RNA replication is governed by membrane vesicles containing pores. The authors used in situ cryo-electron tomography to reveal the minimal constituents of a SARS-CoV-2 “replicopore” and its role in the formation of replication vesicles.
- Liv Zimmermann
- , Xiaohan Zhao
- & Petr Chlanda
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Architecture and regulation of a GDNF-GFRα1 synaptic adhesion assembly
The soluble neurotrophic factor GDNF promotes trans-synaptic adhesion through its co-receptor GFRα1. Here, the authors describe the structural basis for GDNF-GFRα1 adhesion and reconstitute assemblies bridging membranes, demonstrating that binding of either the RET receptor or proteoglycans can disrupt this adhesive function.
- F. M. Houghton
- , S. E. Adams
- & N. Q. McDonald
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Cryo-electron tomography of NLRP3-activated ASC complexes reveals organelle co-localization
The authors characterized puncta in the ASC complex by correlative light microscopy and cryo-ET in cells and propose an ultrastructure of the ASC filament network.
- Yangci Liu
- , Haoming Zhai
- & Yorgo Modis
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Structural insights into the modulation of coronavirus spike tilting and infectivity by hinge glycans
In this work, the authors combine cryo-electron tomography of vitrified virions, mass spectrometry, modeling, molecular dynamics and infectivity assay and report that a hinge glycan on viral spike protein influences virus infection and immune evasion.
- David Chmielewski
- , Eric A. Wilson
- & Wah Chiu
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Hierarchical organization and assembly of the archaeal cell sheath from an amyloid-like protein
Cellular cryoET reveals how an amyloid-like protein of the prototypical archaeon, Methanospirillum hungatei, oligomerizes into a ring containing a giant 2700-strand β sheet, and how rings stack into hoops and into the cylindrical sheath of the cell.
- Hui Wang
- , Jiayan Zhang
- & Z. Hong Zhou
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Streamlined structure determination by cryo-electron tomography and subtomogram averaging using TomoBEAR
Cryo-electron tomography (cryo-ET) enables structural analysis of molecules in situ, but the process is demanding. Here, authors report a software package, TomoBEAR, that streamlines data processing yielding high resolution structures with minimal user input.
- Nikita Balyschew
- , Artsemi Yushkevich
- & Mikhail Kudryashev
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Structure of native chromatin fibres revealed by Cryo-ET in situ
Here, using cryo-FIB and cryo-ET, the authors delineate the architecture of native chromatin fibers and decipher the in situ nucleosomes structure, inspiring future chromatin research.
- Zhen Hou
- , Frank Nightingale
- & Peijun Zhang
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Organisation of the orthobunyavirus tripodal spike and the structural changes induced by low pH and K+ during entry
Enveloped viruses employ the maturing environment of endosomes to promote endosomal escape. Here, authors generate a pseudo-atomic model of the BUNV envelope using sub-tomogram averaging and AlphaFold, and identify ionic cues for fusion events.
- Samantha Hover
- , Frank W. Charlton
- & Juan Fontana
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Intrinsically disordered CsoS2 acts as a general molecular thread for α-carboxysome shell assembly
Carboxysomes are bacterial proteinaceous organelle encapsulating enzymes and pathways to enhance carbon fixation. Here, authors engineer and determine cryoEM structure of minimal α-carboxysome shells to uncover the principle of shell assembly and encapsulation by CsoS2.
- Tao Ni
- , Qiuyao Jiang
- & Peijun Zhang
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FAP106 is an interaction hub for assembling microtubule inner proteins at the cilium inner junction
Microtubule inner proteins (MIPs) contribute to species-specific motility characteristics but are largely unstudied. Here, the authors combine functional, structural and proteomic analysis in T. brucei to advance fundamental understanding of MIP assembly and identify trypanosome-specific MIPs required for motility.
- Michelle M. Shimogawa
- , Angeline S. Wijono
- & Kent L. Hill
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Molecular architecture and conservation of an immature human endogenous retrovirus
The hexagonal immature capsid lattice of human endogenous retrovirus K is determined at 3.2 Å resolution, which is an assembly of small molecule-stabilized hexamers via dimer and trimer interfaces, a highly conserved mechanism among retroviruses.
- Anna-Sophia Krebs
- , Hsuan-Fu Liu
- & Peijun Zhang
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Origin and arrangement of actin filaments for gliding motility in apicomplexan parasites revealed by cryo-electron tomography
Apicomplexan parasites utilize a unique actomyosin system to mediate motility and host cell invasion. Here, the authors apply cryo-ET to Cryptosporidium parvum and Toxoplasma gondii to visualize the F-actin architecture in the native cellular context.
- Matthew Martinez
- , Shrawan Kumar Mageswaran
- & Yi-Wei Chang
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Complement is activated by elevated IgG3 hexameric platforms and deposits C4b onto distinct antibody domains
IgG3 antibodies have potent effector functions, but are not used as therapeutics and structural data are missing. Here, the authors combine cryoEM and MS to study IgG3-mediated complement activation to provide the first structural insights into IgG3.
- Leoni Abendstein
- , Douwe J. Dijkstra
- & Thomas H. Sharp
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Membrane translocation process revealed by in situ structures of type II secretion system secretins
In this work the authors investigate two types of secretins in Escherichia coli, GspDα and GspDβ, and report the Cryo-ET in situ structures of their key intermediate states of during the translocation process. Yielding a resolution ranging from 9 Å to 19 Å, the structures allow the identification of different membrane interaction patterns and ways of transitioning the peptidoglycan layer. These results suggest two distinct models for the membrane translocation of GspDα and GspDβ and provide insights into the inner to outer membrane biogenesis of T2SS secretins.
- Zhili Yu
- , Yaoming Wu
- & Zhao Wang
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A method for restoring signals and revealing individual macromolecule states in cryo-ET, REST
Heavy noise and missing wedge effect hamper the efficient visualization and analysis in cryo-ET. Here, authors present a deep learning-based method for directly visualizing and revealing the dynamic states of target molecules.
- Haonan Zhang
- , Yan Li
- & Ping Zhu
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The in-tissue molecular architecture of β-amyloid pathology in the mammalian brain
In this work the authors reveal in-situ cryoET characterization of β-amyloid plaques in the brain of a mouse model of Alzheimer’s disease.
- Conny Leistner
- , Martin Wilkinson
- & René A. W. Frank
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Cryo-tomography reveals rigid-body motion and organization of apicomplexan invasion machinery
In this study, the authors use cryo-focused-ion-beammilling and cryo-electron tomography to image the apical complex of parasites in their native states. They report insights into the parasite invasion machinery in its protruded and retracted states in three dimensions, including all cytoskeletal assemblies, secretory organelles, and membranes intact.
- Long Gui
- , William J. O’Shaughnessy
- & Daniela Nicastro
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Commercial influenza vaccines vary in HA-complex structure and in induction of cross-reactive HA antibodies
Here, Myers and Gallagher et al. characterize the structural organization of commercial influenza vaccines. The vaccines differ in their structural composition and identify a “spiked nanodisc” arrangement of hemagglutinin (HA) with increased display and immunogenicity of the conserved stem region of HA.
- Mallory L. Myers
- , John R. Gallagher
- & Audray K. Harris
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Determining protein structures in cellular lamella at pseudo-atomic resolution by GisSPA
High-resolution in situ protein structure can be solved by cryo-ET, which requires several days of data collection. Here Cheng et al. report GisSPA, a program that may enable determining sub-4 Å resolution structures on cellular lamellae within one day of data collection.
- Jing Cheng
- , Tong Liu
- & Xinzheng Zhang
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Variable microtubule architecture in the malaria parasite
Microtubules are a ubiquitous eukaryotic cytoskeletal element typically consisting of 13 protofilaments arranged in a hollow cylinder. Using CryoEM and subvolume averaging, Ferreira and Pražák et al. show that Plasmodium does not adhere to a single microtubule structure. Instead, the cytoskeleton changes substantially to produce a unique, fit for purpose structure and organisation at each stage of its life cycle.
- Josie L. Ferreira
- , Vojtěch Pražák
- & Kay Grünewald
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CryoET reveals organelle phenotypes in huntington disease patient iPSC-derived and mouse primary neurons
Huntington’s disease (HD) is a fatal neurodegenerative disorder caused by a genetic mutation in the huntingtin gene (HTT). Here, cryo electron tomography provides insights into the morphology of the cells derived from patients with HD and mouse models of the disease.
- Gong-Her Wu
- , Charlene Smith-Geater
- & Wah Chiu
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Article
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Plasma FIB milling for the determination of structures in situ
The authors harness plasma focused ion beams for pseudo-atomic structure determination, reporting increased throughput and automation in in situ structural biology to elucidate structure-function relationships inside cells and tissues.
- Casper Berger
- , Maud Dumoux
- & Michael Grange
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Article
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Direct Cryo-ET observation of platelet deformation induced by SARS-CoV-2 spike protein
SARS-CoV-2 infection can cause coagulopathic events. Kuhn et al. show by cryo-ET how the Spike protein impacts platelet morphology by binding to the platelet surface, leading to micro-activation; and investigate integrin as possible point of contact.
- Christopher Cyrus Kuhn
- , Nirakar Basnet
- & Naoko Mizuno
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Exocytosis of the silicified cell wall of diatoms involves extensive membrane disintegration
Exocytosis is a fundamental cellular process. Here, the authors report an unusual exocytosis mechanism in the silicified cell wall of diatoms, in which membrane patches are discarded.
- Diede de Haan
- , Lior Aram
- & Assaf Gal
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Article
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Morphological control enables nanometer-scale dissection of cell-cell signaling complexes
Micropatterning on electron microscopy grids enables control of cell positioning and arrangement. Here, the authors describe detailed nanoscale characterization of a micropatterning workflow for reproducible molecular characterization through cryo-ET
- Liam P. Dow
- , Guido Gaietta
- & Dorit Hanein
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Cryo-electron tomography reveals structural insights into the membrane remodeling mode of dynamin-like EHD filaments
Eps15-homology domain containing proteins comprise a family of dynamin-related ATPases. Here, Melo et al. use cryo-electron tomography to determine the membrane-bound EHD4 structure, therefore clarifying the membrane binding and oligomerization mode.
- Arthur A. Melo
- , Thiemo Sprink
- & Oliver Daumke
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Structures of the eukaryotic ribosome and its translational states in situ
The translational states of eukaryotic ribosomes have so far been only investigated in vitro. Here, authors obtained the 3.8 Å in situ 80S ribosome structure, the distribution of translational states and unique arrangement of rRNA expansion segments.
- Patrick C. Hoffmann
- , Jan Philipp Kreysing
- & Martin Beck
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Article
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Donor-strand exchange drives assembly of the TasA scaffold in Bacillus subtilis biofilms
Fibres formed by protein TasA are important components of the extracellular matrix in biofilms developed by the bacterium Bacillus subtilis. Here, Böhning et al. use electron cryomicroscopy and other techniques to show how TasA globular monomers assemble through donor-strand exchange into β-sheet-rich fibres, which in turn assemble into bundles.
- Jan Böhning
- , Mnar Ghrayeb
- & Tanmay A. M. Bharat
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Isotropic reconstruction for electron tomography with deep learning
Cryogenic electron tomography suffers from anisotropic resolution due to the missing-wedge problem. Here, the authors present IsoNet, a neural network that learn the feature representation from similar structures in the tomogram and recover the missing information for isotropic tomogram reconstruction.
- Yun-Tao Liu
- , Heng Zhang
- & Z. Hong Zhou
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Membrane-assisted assembly and selective secretory autophagy of enteroviruses
Enteroviruses are non-enveloped positive-sense RNA viruses that modulate cytoplasmic membranes for replication. To enlighten how enteroviruses assemble around nascent RNA genomes and get package into autophagosomes for release, Dahmane et al. perform cryo-electron tomography of poliovirus-infected cells. They find assembly intermediates that are only present on the cytosolic side of the replication compartment and provide evidence that host factor VPS34 is involved in progression of assembly intermediates.
- Selma Dahmane
- , Adeline Kerviel
- & Lars-Anders Carlson
Ribosomes hibernate on mitochondria during cellular stress