Journal: ACS Nano / Year: 2019 Title: Iron-Sequestering Nanocompartments as Multiplexed Electron Microscopy Gene Reporters. Authors: Felix Sigmund / Susanne Pettinger / Massimo Kube / Fabian Schneider / Martina Schifferer / Steffen Schneider / Maria V Efremova / Jesús Pujol-Martí / Michaela Aichler / Axel Walch / Thomas ...Authors: Felix Sigmund / Susanne Pettinger / Massimo Kube / Fabian Schneider / Martina Schifferer / Steffen Schneider / Maria V Efremova / Jesús Pujol-Martí / Michaela Aichler / Axel Walch / Thomas Misgeld / Hendrik Dietz / 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 ...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 (Qt), which in combination with the recently characterized encapsulin system from (Mx) enables multiplexed gene reporter imaging conventional transmission electron microscopy (TEM) in mammalian cells. Cryo-electron reconstructions revealed that the Qt encapsulin shell self-assembles to nanospheres with = 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 a deep learning classifier to enable automated multiplexed EM gene reporter imaging.
History
Deposition
Apr 18, 2019
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Header (metadata) release
Apr 24, 2019
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Map release
Jun 26, 2019
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Update
Jul 31, 2019
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Current status
Jul 31, 2019
Processing site: PDBe / Status: Released
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