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Updated: Sep 16, 2025
Mitochondria and Endoplasmic Reticulum Imaging by Correlative Light and Volume Electron Microscopy
Published on: July 20, 2019
Super-resolution microscopy of mitochondrial mRNAs
Stefan Stoldt1,2,3, Frederike Maass1,2,3, Michael Weber4
1Department of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, RG Mitochondrial Structure and Dynamics, Göttingen, Germany.
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.
Insights
Researchers visualized individual mitochondrial messenger RNA (mRNA) molecules using super-resolution microscopy. This technique reveals how mRNA distribution and shape change in cells, offering new insights into mitochondrial gene regulation.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- Mitochondria possess their own DNA (mtDNA) and gene expression machinery.
- The spatial distribution of mitochondrial mRNA and proteins is poorly understood due to microscopy limitations.
Purpose of the Study:
- To develop and apply super-resolution microscopy techniques to visualize individual mitochondrial mRNA molecules and associated proteins.
- To investigate the spatial organization and dynamics of mitochondrial mRNA in response to cellular stress and disease.
Main Methods:
- Single-molecule fluorescence in situ hybridization (smFISH) combined with STED and MINFLUX super-resolution microscopy.
- Visualization of individual mitochondrial mRNA molecules and RNA-binding proteins.
- Analysis of mRNA distribution, shape, and proximity to ribosomes in mammalian cells.
Main Results:
- STED nanoscopy revealed spatial relationships between different mRNA species and proteins like GRSF1.
- Adaptive changes in mRNA distribution and quantity were observed in stressed cells and patient-derived cell lines.
- STED-smFISH showed mRNA release during apoptosis, and MINFLUX revealed mRNA folding and proximity to mitochondrial ribosomes.
Conclusions:
- Super-resolution microscopy enables visualization of mitochondrial mRNA and associated proteins at the single-molecule level.
- Mitochondrial mRNA distribution and organization are dynamic and adapt to cellular conditions, including apoptosis.
- These advanced imaging techniques provide new avenues for studying mitochondrial gene regulation in health and disease.
