Super-resolution microscopy of mitochondrial mRNAs
Abstract Mitochondria contain their own DNA (mtDNA) and a dedicated gene expression machinery. As the mitochondrial dimensions are close to the diffraction limit of classical light microscopy, the spatial distribution of mitochondrial proteins and in particular of mitochondrial mRNAs remains underex...
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2025-07-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-025-61577-5 |
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| author | Stefan Stoldt Frederike Maass Michael Weber Sven Dennerlein Peter Ilgen Jutta Gärtner Aysenur Canfes Sarah V. Schweighofer Daniel C. Jans Peter Rehling Stefan Jakobs |
| author_facet | Stefan Stoldt Frederike Maass Michael Weber Sven Dennerlein Peter Ilgen Jutta Gärtner Aysenur Canfes Sarah V. Schweighofer Daniel C. Jans Peter Rehling Stefan Jakobs |
| author_sort | Stefan Stoldt |
| collection | DOAJ |
| description | Abstract Mitochondria contain their own DNA (mtDNA) and a dedicated gene expression machinery. As the mitochondrial dimensions are close to the diffraction limit of classical light microscopy, the spatial distribution of mitochondrial proteins and in particular of mitochondrial mRNAs remains underexplored. Here, we establish single-molecule fluorescence in situ hybridization (smFISH) combined with STED and MINFLUX super-resolution microscopy (nanoscopy) to visualize individual mitochondrial mRNA molecules and associated proteins. STED nanoscopy reveals the spatial relationships between distinct mRNA species and proteins such as the RNA granule marker GRSF1, demonstrating adaptive changes in mRNA distribution and quantity in challenged mammalian cells and patient-derived cell lines. Notably, STED-smFISH shows the release of mRNAs during apoptosis, while MINFLUX reveals the folding of the mRNAs into variable shapes, as well as their spatial proximity to mitochondrial ribosomes. These protocols are transferable to various cell types and open new avenues for understanding mitochondrial gene regulation in health and disease. |
| format | Article |
| id | doaj-art-2d85e720fd784d9e93507c0d285b5e5d |
| institution | DOAJ |
| issn | 2041-1723 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj-art-2d85e720fd784d9e93507c0d285b5e5d2025-08-20T03:05:14ZengNature PortfolioNature Communications2041-17232025-07-0116111110.1038/s41467-025-61577-5Super-resolution microscopy of mitochondrial mRNAsStefan Stoldt0Frederike Maass1Michael Weber2Sven Dennerlein3Peter Ilgen4Jutta Gärtner5Aysenur Canfes6Sarah V. Schweighofer7Daniel C. Jans8Peter Rehling9Stefan Jakobs10Department of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary SciencesDepartment of Cellular Biochemistry, University Medical Center GöttingenDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsCluster of Excellence “Multiscale Bioimaging: from Molecular Machines to Networks of Excitable Cells” (MBExC), University of GöttingenDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsCluster of Excellence “Multiscale Bioimaging: from Molecular Machines to Networks of Excitable Cells” (MBExC), University of GöttingenDepartment of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and DynamicsAbstract Mitochondria contain their own DNA (mtDNA) and a dedicated gene expression machinery. As the mitochondrial dimensions are close to the diffraction limit of classical light microscopy, the spatial distribution of mitochondrial proteins and in particular of mitochondrial mRNAs remains underexplored. Here, we establish single-molecule fluorescence in situ hybridization (smFISH) combined with STED and MINFLUX super-resolution microscopy (nanoscopy) to visualize individual mitochondrial mRNA molecules and associated proteins. STED nanoscopy reveals the spatial relationships between distinct mRNA species and proteins such as the RNA granule marker GRSF1, demonstrating adaptive changes in mRNA distribution and quantity in challenged mammalian cells and patient-derived cell lines. Notably, STED-smFISH shows the release of mRNAs during apoptosis, while MINFLUX reveals the folding of the mRNAs into variable shapes, as well as their spatial proximity to mitochondrial ribosomes. These protocols are transferable to various cell types and open new avenues for understanding mitochondrial gene regulation in health and disease.https://doi.org/10.1038/s41467-025-61577-5 |
| spellingShingle | Stefan Stoldt Frederike Maass Michael Weber Sven Dennerlein Peter Ilgen Jutta Gärtner Aysenur Canfes Sarah V. Schweighofer Daniel C. Jans Peter Rehling Stefan Jakobs Super-resolution microscopy of mitochondrial mRNAs Nature Communications |
| title | Super-resolution microscopy of mitochondrial mRNAs |
| title_full | Super-resolution microscopy of mitochondrial mRNAs |
| title_fullStr | Super-resolution microscopy of mitochondrial mRNAs |
| title_full_unstemmed | Super-resolution microscopy of mitochondrial mRNAs |
| title_short | Super-resolution microscopy of mitochondrial mRNAs |
| title_sort | super resolution microscopy of mitochondrial mrnas |
| url | https://doi.org/10.1038/s41467-025-61577-5 |
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