TY - JOUR A1 - Sperber, Hannah Sabeth A1 - Welke, Robert-William A1 - Petazzi, Roberto Arturo A1 - Bergmann, Ronny A1 - Schade, Matthias A1 - Shai, Yechiel A1 - Chiantia, Salvatore A1 - Herrmann, Andreas A1 - Schwarzer, Roland T1 - Self-association and subcellular localization of Puumala hantavirus envelope proteins JF - Scientific reports N2 - Hantavirus assembly and budding are governed by the surface glycoproteins Gn and Gc. In this study, we investigated the glycoproteins of Puumala, the most abundant Hantavirus species in Europe, using fluorescently labeled wild-type constructs and cytoplasmic tail (CT) mutants. We analyzed their intracellular distribution, co-localization and oligomerization, applying comprehensive live, single-cell fluorescence techniques, including confocal microscopy, imaging flow cytometry, anisotropy imaging and Number&Brightness analysis. We demonstrate that Gc is significantly enriched in the Golgi apparatus in absence of other viral components, while Gn is mainly restricted to the endoplasmic reticulum (ER). Importantly, upon co-expression both glycoproteins were found in the Golgi apparatus. Furthermore, we show that an intact CT of Gc is necessary for efficient Golgi localization, while the CT of Gn influences protein stability. Finally, we found that Gn assembles into higher-order homo-oligomers, mainly dimers and tetramers, in the ER while Gc was present as mixture of monomers and dimers within the Golgi apparatus. Our findings suggest that PUUV Gc is the driving factor of the targeting of Gc and Gn to the Golgi region, while Gn possesses a significantly stronger self-association potential. Y1 - 2019 U6 - https://doi.org/10.1038/s41598-018-36879-y SN - 2045-2322 VL - 9 PB - Nature Publ. Group CY - London ER - TY - GEN A1 - Sperber, Hannah Sabeth A1 - Welke, Robert-William A1 - Petazzi, Roberto Arturo A1 - Bergmann, Ronny A1 - Schade, Matthias A1 - Shai, Yechiel A1 - Chiantia, Salvatore A1 - Herrmann, Andreas A1 - Schwarzer, Roland T1 - Self-association and subcellular localization of Puumala hantavirus envelope proteins T2 - Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Hantavirus assembly and budding are governed by the surface glycoproteins Gn and Gc. In this study, we investigated the glycoproteins of Puumala, the most abundant Hantavirus species in Europe, using fluorescently labeled wild-type constructs and cytoplasmic tail (CT) mutants. We analyzed their intracellular distribution, co-localization and oligomerization, applying comprehensive live, single-cell fluorescence techniques, including confocal microscopy, imaging flow cytometry, anisotropy imaging and Number&Brightness analysis. We demonstrate that Gc is significantly enriched in the Golgi apparatus in absence of other viral components, while Gn is mainly restricted to the endoplasmic reticulum (ER). Importantly, upon co-expression both glycoproteins were found in the Golgi apparatus. Furthermore, we show that an intact CT of Gc is necessary for efficient Golgi localization, while the CT of Gn influences protein stability. Finally, we found that Gn assembles into higher-order homo-oligomers, mainly dimers and tetramers, in the ER while Gc was present as mixture of monomers and dimers within the Golgi apparatus. Our findings suggest that PUUV Gc is the driving factor of the targeting of Gc and Gn to the Golgi region, while Gn possesses a significantly stronger self-association potential. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 648 KW - Sin-Nombre-Virus KW - nucleocapsid protein KW - cytoplasmic tails KW - electron cryotomography KW - autophagic clearance KW - glycoprotein KW - Gn KW - G1 KW - brightness KW - fever Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-425040 SN - 1866-8372 IS - 648 ER - TY - JOUR A1 - Welke, Robert-William A1 - Sperber, Hannah Sabeth A1 - Bergmann, Ronny A1 - Koikkarah, Amit A1 - Menke, Laura A1 - Sieben, Christian A1 - Krüger, Detlev H. A1 - Chiantia, Salvatore A1 - Herrmann, Andreas A1 - Schwarzer, Roland T1 - Characterization of hantavirus N protein intracellular dynamics and localization JF - Viruses N2 - Hantaviruses are enveloped viruses that possess a tri-segmented, negative-sense RNA genome. The viral S-segment encodes the multifunctional nucleocapsid protein (N), which is involved in genome packaging, intracellular protein transport, immunoregulation, and several other crucial processes during hantavirus infection. In this study, we generated fluorescently tagged N protein constructs derived from Puumalavirus (PUUV), the dominant hantavirus species in Central, Northern, and Eastern Europe. We comprehensively characterized this protein in the rodent cell line CHO-K1, monitoring the dynamics of N protein complex formation and investigating co-localization with host proteins as well as the viral glycoproteins Gc and Gn. We observed formation of large, fibrillar PUUV N protein aggregates, rapidly coalescing from early punctate and spike-like assemblies. Moreover, we found significant spatial correlation of N with vimentin, actin, and P-bodies but not with microtubules. N constructs also co-localized with Gn and Gc albeit not as strongly as the glycoproteins associated with each other. Finally, we assessed oligomerization of N constructs, observing efficient and concentration-dependent multimerization, with complexes comprising more than 10 individual proteins. KW - hantavirus KW - N protein KW - oligomerization KW - actin KW - P-bodies KW - vimentin KW - Number and Brightness KW - Puumalavirus KW - macromolecular assemblies Y1 - 2022 U6 - https://doi.org/10.3390/v14030457 SN - 1999-4915 VL - 14 IS - 3 PB - MDPI CY - Basel ER -