Cargando…

Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells

Imaging of actin filaments is crucial due to the integral role that they play in many cellular functions such as intracellular transport, membrane remodelling and cell motility. Visualizing actin filaments has so far relied on fluorescence microscopy and electron microscopy/tomography. The former la...

Descripción completa

Detalles Bibliográficos
Autores principales: Koronfel, Mohamed, Kounatidis, Ilias, Mwangangi, Dennis M., Vyas, Nina, Okolo, Chidinma, Jadhav, Archana, Fish, Tom, Chotchuang, Phatcharin, Schulte, Albert, Robinson, Robert C., Harkiolaki, Maria
Formato: Online Artículo Texto
Lenguaje:English
Publicado: International Union of Crystallography 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8647181/
https://www.ncbi.nlm.nih.gov/pubmed/34866605
http://dx.doi.org/10.1107/S2059798321010329
_version_ 1784610563372875776
author Koronfel, Mohamed
Kounatidis, Ilias
Mwangangi, Dennis M.
Vyas, Nina
Okolo, Chidinma
Jadhav, Archana
Fish, Tom
Chotchuang, Phatcharin
Schulte, Albert
Robinson, Robert C.
Harkiolaki, Maria
author_facet Koronfel, Mohamed
Kounatidis, Ilias
Mwangangi, Dennis M.
Vyas, Nina
Okolo, Chidinma
Jadhav, Archana
Fish, Tom
Chotchuang, Phatcharin
Schulte, Albert
Robinson, Robert C.
Harkiolaki, Maria
author_sort Koronfel, Mohamed
collection PubMed
description Imaging of actin filaments is crucial due to the integral role that they play in many cellular functions such as intracellular transport, membrane remodelling and cell motility. Visualizing actin filaments has so far relied on fluorescence microscopy and electron microscopy/tomography. The former lacks the capacity to capture the overall local ultrastructure, while the latter requires rigorous sample preparation that can lead to potential artefacts, and only delivers relatively small volumes of imaging data at the thinnest areas of a cell. In this work, a correlative approach utilizing in situ super-resolution fluorescence imaging and cryo X-ray tomography was used to image bundles of actin filaments deep inside cells under near-native conditions. In this case, fluorescence 3D imaging localized the actin bundles within the intracellular space, while X-ray tomograms of the same areas provided detailed views of the local ultrastructure. Using this new approach, actin trails connecting vesicles in the perinuclear area and hotspots of actin presence within and around multivesicular bodies were observed. The characteristic prevalence of filamentous actin in cytoplasmic extensions was also documented.
format Online
Article
Text
id pubmed-8647181
institution National Center for Biotechnology Information
language English
publishDate 2021
publisher International Union of Crystallography
record_format MEDLINE/PubMed
spelling pubmed-86471812021-12-16 Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells Koronfel, Mohamed Kounatidis, Ilias Mwangangi, Dennis M. Vyas, Nina Okolo, Chidinma Jadhav, Archana Fish, Tom Chotchuang, Phatcharin Schulte, Albert Robinson, Robert C. Harkiolaki, Maria Acta Crystallogr D Struct Biol Ccp4 Imaging of actin filaments is crucial due to the integral role that they play in many cellular functions such as intracellular transport, membrane remodelling and cell motility. Visualizing actin filaments has so far relied on fluorescence microscopy and electron microscopy/tomography. The former lacks the capacity to capture the overall local ultrastructure, while the latter requires rigorous sample preparation that can lead to potential artefacts, and only delivers relatively small volumes of imaging data at the thinnest areas of a cell. In this work, a correlative approach utilizing in situ super-resolution fluorescence imaging and cryo X-ray tomography was used to image bundles of actin filaments deep inside cells under near-native conditions. In this case, fluorescence 3D imaging localized the actin bundles within the intracellular space, while X-ray tomograms of the same areas provided detailed views of the local ultrastructure. Using this new approach, actin trails connecting vesicles in the perinuclear area and hotspots of actin presence within and around multivesicular bodies were observed. The characteristic prevalence of filamentous actin in cytoplasmic extensions was also documented. International Union of Crystallography 2021-11-29 /pmc/articles/PMC8647181/ /pubmed/34866605 http://dx.doi.org/10.1107/S2059798321010329 Text en © Mohamed Koronfel et al. 2021 https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution (CC-BY) Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.
spellingShingle Ccp4
Koronfel, Mohamed
Kounatidis, Ilias
Mwangangi, Dennis M.
Vyas, Nina
Okolo, Chidinma
Jadhav, Archana
Fish, Tom
Chotchuang, Phatcharin
Schulte, Albert
Robinson, Robert C.
Harkiolaki, Maria
Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells
title Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells
title_full Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells
title_fullStr Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells
title_full_unstemmed Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells
title_short Correlative cryo-imaging of the cellular universe with soft X-rays and laser light used to track F-actin structures in mammalian cells
title_sort correlative cryo-imaging of the cellular universe with soft x-rays and laser light used to track f-actin structures in mammalian cells
topic Ccp4
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8647181/
https://www.ncbi.nlm.nih.gov/pubmed/34866605
http://dx.doi.org/10.1107/S2059798321010329
work_keys_str_mv AT koronfelmohamed correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT kounatidisilias correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT mwangangidennism correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT vyasnina correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT okolochidinma correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT jadhavarchana correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT fishtom correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT chotchuangphatcharin correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT schultealbert correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT robinsonrobertc correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells
AT harkiolakimaria correlativecryoimagingofthecellularuniversewithsoftxraysandlaserlightusedtotrackfactinstructuresinmammaliancells