---
_id: '12239'
abstract:
- lang: eng
  text: Biological systems are the sum of their dynamic three-dimensional (3D) parts.
    Therefore, it is critical to study biological structures in 3D and at high resolution
    to gain insights into their physiological functions. Electron microscopy of metal
    replicas of unroofed cells and isolated organelles has been a key technique to
    visualize intracellular structures at nanometer resolution. However, many of these
    methods require specialized equipment and personnel to complete them. Here, we
    present novel accessible methods to analyze biological structures in unroofed
    cells and biochemically isolated organelles in 3D and at nanometer resolution,
    focusing on Arabidopsis clathrin-coated vesicles (CCVs). While CCVs are essential
    trafficking organelles, their detailed structural information is lacking due to
    their poor preservation when observed via classical electron microscopy protocols
    experiments. First, we establish a method to visualize CCVs in unroofed cells
    using scanning transmission electron microscopy tomography, providing sufficient
    resolution to define the clathrin coat arrangements. Critically, the samples are
    prepared directly on electron microscopy grids, removing the requirement to use
    extremely corrosive acids, thereby enabling the use of this method in any electron
    microscopy lab. Secondly, we demonstrate that this standardized sample preparation
    allows the direct comparison of isolated CCV samples with those visualized in
    cells. Finally, to facilitate the high-throughput and robust screening of metal
    replicated samples, we provide a deep learning analysis method to screen the “pseudo
    3D” morphologies of CCVs imaged with 2D modalities. Collectively, our work establishes
    accessible ways to examine the 3D structure of biological samples and provide
    novel insights into the structure of plant CCVs.
acknowledged_ssus:
- _id: EM-Fac
- _id: LifeSc
- _id: Bio
acknowledgement: A.J. is supported by funding from the Austrian Science Fund I3630B25
  (to J.F.). This research was supported by the Scientific Service Units of Institute
  of Science and Technology Austria (ISTA) through resources provided by the Electron
  Microscopy Facility, Lab Support Facility, and the Imaging and Optics Facility.
  We acknowledge Prof. David Robinson (Heidelberg) and Prof. Jan Traas (Lyon) for
  making us aware of previously published classical on-grid preparation methods. No
  conflict of interest declared.
article_processing_charge: Yes (via OA deal)
article_type: original
author:
- first_name: Alexander J
  full_name: Johnson, Alexander J
  id: 46A62C3A-F248-11E8-B48F-1D18A9856A87
  last_name: Johnson
  orcid: 0000-0002-2739-8843
- first_name: Walter
  full_name: Kaufmann, Walter
  id: 3F99E422-F248-11E8-B48F-1D18A9856A87
  last_name: Kaufmann
  orcid: 0000-0001-9735-5315
- first_name: Christoph M
  full_name: Sommer, Christoph M
  id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87
  last_name: Sommer
  orcid: 0000-0003-1216-9105
- first_name: Tommaso
  full_name: Costanzo, Tommaso
  id: D93824F4-D9BA-11E9-BB12-F207E6697425
  last_name: Costanzo
  orcid: 0000-0001-9732-3815
- first_name: Dana A.
  full_name: Dahhan, Dana A.
  last_name: Dahhan
- first_name: Sebastian Y.
  full_name: Bednarek, Sebastian Y.
  last_name: Bednarek
- first_name: Jiří
  full_name: Friml, Jiří
  id: 4159519E-F248-11E8-B48F-1D18A9856A87
  last_name: Friml
  orcid: 0000-0002-8302-7596
citation:
  ama: Johnson AJ, Kaufmann W, Sommer CM, et al. Three-dimensional visualization of
    planta clathrin-coated vesicles at ultrastructural resolution. <i>Molecular Plant</i>.
    2022;15(10):1533-1542. doi:<a href="https://doi.org/10.1016/j.molp.2022.09.003">10.1016/j.molp.2022.09.003</a>
  apa: Johnson, A. J., Kaufmann, W., Sommer, C. M., Costanzo, T., Dahhan, D. A., Bednarek,
    S. Y., &#38; Friml, J. (2022). Three-dimensional visualization of planta clathrin-coated
    vesicles at ultrastructural resolution. <i>Molecular Plant</i>. Elsevier. <a href="https://doi.org/10.1016/j.molp.2022.09.003">https://doi.org/10.1016/j.molp.2022.09.003</a>
  chicago: Johnson, Alexander J, Walter Kaufmann, Christoph M Sommer, Tommaso Costanzo,
    Dana A. Dahhan, Sebastian Y. Bednarek, and Jiří Friml. “Three-Dimensional Visualization
    of Planta Clathrin-Coated Vesicles at Ultrastructural Resolution.” <i>Molecular
    Plant</i>. Elsevier, 2022. <a href="https://doi.org/10.1016/j.molp.2022.09.003">https://doi.org/10.1016/j.molp.2022.09.003</a>.
  ieee: A. J. Johnson <i>et al.</i>, “Three-dimensional visualization of planta clathrin-coated
    vesicles at ultrastructural resolution,” <i>Molecular Plant</i>, vol. 15, no.
    10. Elsevier, pp. 1533–1542, 2022.
  ista: Johnson AJ, Kaufmann W, Sommer CM, Costanzo T, Dahhan DA, Bednarek SY, Friml
    J. 2022. Three-dimensional visualization of planta clathrin-coated vesicles at
    ultrastructural resolution. Molecular Plant. 15(10), 1533–1542.
  mla: Johnson, Alexander J., et al. “Three-Dimensional Visualization of Planta Clathrin-Coated
    Vesicles at Ultrastructural Resolution.” <i>Molecular Plant</i>, vol. 15, no.
    10, Elsevier, 2022, pp. 1533–42, doi:<a href="https://doi.org/10.1016/j.molp.2022.09.003">10.1016/j.molp.2022.09.003</a>.
  short: A.J. Johnson, W. Kaufmann, C.M. Sommer, T. Costanzo, D.A. Dahhan, S.Y. Bednarek,
    J. Friml, Molecular Plant 15 (2022) 1533–1542.
corr_author: '1'
date_created: 2023-01-16T09:51:49Z
date_published: 2022-10-03T00:00:00Z
date_updated: 2025-04-15T07:32:09Z
day: '03'
ddc:
- '580'
department:
- _id: JiFr
- _id: EM-Fac
- _id: Bio
doi: 10.1016/j.molp.2022.09.003
external_id:
  isi:
  - '000882769800009'
  pmid:
  - '36081349'
file:
- access_level: open_access
  checksum: 04d5c12490052d03e4dc4412338a43dd
  content_type: application/pdf
  creator: dernst
  date_created: 2023-01-30T07:46:51Z
  date_updated: 2023-01-30T07:46:51Z
  file_id: '12435'
  file_name: 2022_MolecularPlant_Johnson.pdf
  file_size: 2307251
  relation: main_file
  success: 1
file_date_updated: 2023-01-30T07:46:51Z
has_accepted_license: '1'
intvolume: '        15'
isi: 1
issue: '10'
keyword:
- Plant Science
- Molecular Biology
language:
- iso: eng
license: https://creativecommons.org/licenses/by/4.0/
month: '10'
oa: 1
oa_version: Published Version
page: 1533-1542
pmid: 1
project:
- _id: 26538374-B435-11E9-9278-68D0E5697425
  call_identifier: FWF
  grant_number: I03630
  name: Molecular mechanisms of endocytic cargo recognition in plants
publication: Molecular Plant
publication_identifier:
  issn:
  - 1674-2052
publication_status: published
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Three-dimensional visualization of planta clathrin-coated vesicles at ultrastructural
  resolution
tmp:
  image: /images/cc_by.png
  legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
  name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
  short: CC BY (4.0)
type: journal_article
user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8
volume: 15
year: '2022'
...