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  Multicolor 3D MINFLUX nanoscopy of mitochondrial MICOS proteins

Pape, J. K., Stephan, T., Balzarotti, F., Büchner, R., Lange, F., Riedel, D., et al. (2020). Multicolor 3D MINFLUX nanoscopy of mitochondrial MICOS proteins. Proceedings of the National Academy of Sciences of the USA, 117(34), 20607-20614. doi:10.1073/pnas.2009364117.

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Item Permalink: http://hdl.handle.net/21.11116/0000-0007-1FF4-4 Version Permalink: http://hdl.handle.net/21.11116/0000-0007-2000-4
Genre: Journal Article

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 Creators:
Pape, J. K.1, Author              
Stephan, T.2, Author              
Balzarotti, F.1, Author              
Büchner, R., Author
Lange, F.3, Author              
Riedel, D.4, Author              
Jakobs, S.3, Author              
Hell, S. W.1, Author              
Affiliations:
1Department of NanoBiophotonics, MPI for Biophysical Chemistry, Max Planck Society, ou_578627              
2Research Group of Mitochondrial Structure and Dynamics, MPI for Biophysical Chemistry, Max Planck Society, ou_578566              
3Research Group of Mitochondrial Structure and Dynamics, MPI for biophysical chemistry, Max Planck Society, ou_578566              
4Facility for Electron Microscopy, MPI for biophysical chemistry, Max Planck Society, ou_578615              

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Free keywords: MINFLUX; superresolution microscopy; mitochondria; MICOS; cluster analysis
 Abstract: The mitochondrial contact site and cristae organizing system (MICOS) is a multisubunit protein complex that is essential for the proper architecture of the mitochondrial inner membrane. MICOS plays a key role in establishing and maintaining crista junctions, tubular or slit-like structures that connect the cristae membrane with the inner boundary membrane, thereby ensuring a contiguous inner membrane. MICOS is enriched at crista junctions, but the detailed distribution of its subunits around crista junctions is unclear because such small length scales are inaccessible with established fluorescence microscopy. By targeting individually activated fluorophores with an excitation beam featuring a central zero-intensity point, the nanoscopy method called MINFLUX delivers single-digit nanometer-scale three-dimensional (3D) resolution and localization precision. We employed MINFLUX nanoscopy to investigate the submitochondrial localization of the core MICOS subunit Mic60 in relation to two other MICOS proteins, Mic10 and Mic19. We demonstrate that dual-color 3D MINFLUX nanoscopy is applicable to the imaging of organellar substructures, yielding a 3D localization precision of ∼5 nm in human mitochondria. This isotropic precision facilitated the development of an analysis framework that assigns localization clouds to individual molecules, thus eliminating a source of bias when drawing quantitative conclusions from single-molecule localization microscopy data. MINFLUX recordings of Mic60 indicate ringlike arrangements of multiple molecules with a diameter of 40 to 50 nm, suggesting that Mic60 surrounds individual crista junctions. Statistical analysis of dual-color MINFLUX images demonstrates that Mic19 is generally in close proximity to Mic60, whereas the spatial coordination of Mic10 with Mic60 is less regular, suggesting structural heterogeneity of MICOS.

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Language(s): eng - English
 Dates: 2020-08-112020-08-25
 Publication Status: Published in print
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 Rev. Type: Peer
 Identifiers: DOI: 10.1073/pnas.2009364117
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Title: Proceedings of the National Academy of Sciences of the USA
Source Genre: Journal
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Pages: - Volume / Issue: 117 (34) Sequence Number: - Start / End Page: 20607 - 20614 Identifier: -