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Sigmund, F. ; Pettinger, S. ; Kube, M.* ; Schneider, F.* ; Schifferer, M.* ; Schneider, S.* ; Efremova, M.V. ; Pujol-Martí, J.* ; Aichler, M. ; Walch, A.K. ; Misgeld, T.* ; Dietz, H.* ; Westmeyer, G.G.

Iron-sequestering nanocompartments as multiplexed Electron Microscopy gene reporters.

ACS Nano 13, 8114-8123 (2019)
Verlagsversion Postprint Forschungsdaten DOI
Open Access Green
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 similar to 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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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Schlagwörter Encapsulin ; Electron Microscopy ; Cryo-electron Microscopy ; Gene Reporter ; Multiplexing ; Compartmentalization ; Iron Biomineralization; Molecular Sociology; Proteins; Ferritin; Membrane; Sites; Label
ISSN (print) / ISBN 1936-0851
e-ISSN 1936-086X
Zeitschrift ACS Nano
Quellenangaben Band: 13, Heft: 7, Seiten: 8114-8123 Artikelnummer: , Supplement: ,
Verlag American Chemical Society (ACS)
Verlagsort 1155 16th St, Nw, Washington, Dc 20036 Usa
Begutachtungsstatus