| First Authors | Christian Franke |
|---|---|
| Authors | Christian Franke, Urska Repnik, Sandra Segeletz, Nicolas Brouilly, Yannis Kalaidzidis, Jean-Marc Verbavatz, Marino Zerial |
| Corresponding Authors | Marino Zerial |
| Last Authors | Marino Zerial |
| Journal Name | Traffic (Copenhagen, Denmark) (Traffic) |
| Volume | 20 |
| Issue | 8 |
| Page Range | 601-617 |
| Open Access | true |
| Print Publication Date | 2019-08-01 |
| Online Publication Date | |
| Abstract | Many cellular organelles, including endosomes, show compartmentalization into distinct functional domains, which, however, cannot be resolved by diffraction-limited light microscopy. Single molecule localization microscopy (SMLM) offers nanoscale resolution but data interpretation is often inconclusive when the ultrastructural context is missing. Correlative light electron microscopy (CLEM) combining SMLM with electron microscopy (EM) enables correlation of functional subdomains of organelles in relation to their underlying ultrastructure at nanometer resolution. However, the specific demands for EM sample preparation and the requirements for fluorescent single-molecule photo-switching are opposed. Here, we developed a novel superCLEM workflow that combines triple-color SMLM (dSTORM & PALM) and electron tomography using semi-thin Tokuyasu thawed cryosections. We applied the superCLEM approach to directly visualize nanoscale compartmentalization of endosomes in HeLa cells. Internalized, fluorescently labeled Transferrin and EGF were resolved into morphologically distinct domains within the same endosome. We found that the small GTPase Rab5 is organized in nanodomains on the globular part of early endosomes. The simultaneous visualization of several proteins in functionally distinct endosomal sub-compartments demonstrates the potential of superCLEM to link the ultrastructure of organelles with their molecular organization at nanoscale resolution. |
| Cover Image | |
| Affiliated With | EM Facility, Postdoc first author, Postdocs, Zerial, Postdoc first male |
| Selected By | |
| Acknowledged Services | Transgenic Core Facility, EM Facility, Technology Development Studio TDS, Genome Engineering Facility, Light Microscopy Facility, FACS Facility |
| Publication Status | Published |
| Edoc Link | |
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| DOI | 10.1111/tra.12671 |
| PubMed ID | 31206952 |
| WebOfScience Link | WOS:000477759000005 |
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| Display Publisher Download Only | false |
| Visible On MPI-CBG Website | true |
| PDF Downloadable | true |
| Created By | thuem |
| Added Date | 2019-07-30 |
| Last Edited By | herbst |
| Last Edited Date | 2021-04-29 17:20:39.16 |
| Library ID | 7440 |
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| Entry Complete | true |
| eDoc Compliant | true |
| Include in Edoc Report | true |
| In Pure | false |
| Ready for eDoc Export | false |
| Author Affiliations Complete | false |
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