Iron-Sequestering Nanocompartments as Multiplexed Electron Microscopy Gene Reporters
- Felix SigmundFelix SigmundDepartment of Nuclear Medicine, TUM School of Medicine, Technical University of Munich, 81675 Munich, GermanyInstitute of Biological and Medical Imaging, Helmholtz Zentrum München, 85764 Neuherberg, GermanyInstitute of Developmental Genetics, Helmholtz Zentrum München, 85764 Neuherberg, GermanyMore by Felix Sigmund
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- Susanne PettingerSusanne PettingerDepartment of Nuclear Medicine, TUM School of Medicine, Technical University of Munich, 81675 Munich, GermanyInstitute of Biological and Medical Imaging, Helmholtz Zentrum München, 85764 Neuherberg, GermanyInstitute of Developmental Genetics, Helmholtz Zentrum München, 85764 Neuherberg, GermanyMore by Susanne Pettinger
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- Massimo KubeMassimo KubeLaboratory for Biomolecular Design, Department of Physics, Technical University of Munich, 85748 Garching, GermanyMore by Massimo Kube
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- Fabian SchneiderFabian SchneiderLaboratory for Biomolecular Design, Department of Physics, Technical University of Munich, 85748 Garching, GermanyMore by Fabian Schneider
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- Martina SchiffererMartina SchiffererInstitute of Neuronal Cell Biology, TUM School of Medicine, Technical University of Munich, 80802 Munich, GermanyGerman Center for Neurodegenerative Diseases (DZNE), 81377 Munich, GermanyMore by Martina Schifferer
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- Steffen SchneiderSteffen SchneiderComputational Neuroengineering, Department of Electrical and Computer Engineering, Technical University of Munich, 80333 Munich, GermanyTübingen AI Center, University of Tübingen, 72076 Tübingen, GermanyMore by Steffen Schneider
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- Maria V. EfremovaMaria V. EfremovaDepartment of Nuclear Medicine, TUM School of Medicine, Technical University of Munich, 81675 Munich, GermanyInstitute of Biological and Medical Imaging, Helmholtz Zentrum München, 85764 Neuherberg, GermanyInstitute of Developmental Genetics, Helmholtz Zentrum München, 85764 Neuherberg, GermanyLaboratory of Chemical Design of Bionanomaterials for Medical Applications, Department of Chemistry, Lomonosov Moscow State University, 119991 Moscow, Russian FederationMore by Maria V. Efremova
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- Jesús Pujol-MartíJesús Pujol-MartíDepartment “Circuits - Computation - Models”, Max Planck Institute of Neurobiology, 82152 Martinsried, GermanyMore by Jesús Pujol-Martí
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- Michaela AichlerMichaela AichlerResearch Unit Analytical Pathology, Helmholtz Zentrum München, 85764 Neuherberg, GermanyMore by Michaela Aichler
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- Axel WalchAxel WalchResearch Unit Analytical Pathology, Helmholtz Zentrum München, 85764 Neuherberg, GermanyMore by Axel Walch
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- Thomas MisgeldThomas MisgeldInstitute of Neuronal Cell Biology, TUM School of Medicine, Technical University of Munich, 80802 Munich, GermanyGerman Center for Neurodegenerative Diseases (DZNE), 81377 Munich, GermanyMore by Thomas Misgeld
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- Hendrik DietzHendrik DietzLaboratory for Biomolecular Design, Department of Physics, Technical University of Munich, 85748 Garching, GermanyMore by Hendrik Dietz
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- Gil G. Westmeyer*Gil G. Westmeyer*E-mail: [email protected]Department of Nuclear Medicine, TUM School of Medicine, Technical University of Munich, 81675 Munich, GermanyInstitute of Biological and Medical Imaging, Helmholtz Zentrum München, 85764 Neuherberg, GermanyInstitute of Developmental Genetics, Helmholtz Zentrum München, 85764 Neuherberg, GermanyMore by Gil G. Westmeyer
Abstract
Multicolored gene reporters for light microscopy are indispensable for biomedical research, but equivalent genetic tools for electron microscopy (EM) are still rare despite the increasing importance of nanometer resolution for reverse engineering of molecular machinery and reliable mapping of cellular circuits. We here introduce the fully genetic encapsulin/cargo system of Quasibacillus thermotolerans (Qt), which in combination with the recently characterized encapsulin system from Myxococcus xanthus (Mx) enables multiplexed gene reporter imaging via conventional transmission electron microscopy (TEM) in mammalian cells. Cryo-electron reconstructions revealed that the Qt encapsulin shell self-assembles to nanospheres with T = 4 icosahedral symmetry and a diameter of ∼43 nm harboring two putative pore regions at the 5-fold and 3-fold axes. We also found that upon heterologous expression in mammalian cells, the native cargo is autotargeted to the inner surface of the shell and exhibits ferroxidase activity leading to efficient intraluminal iron biomineralization, which enhances cellular TEM contrast. We furthermore demonstrate that the two differently sized encapsulins of Qt and Mx do not intermix and can be robustly differentiated by conventional TEM via a deep learning classifier to enable automated multiplexed EM gene reporter imaging.
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