Zika virus replication in glioblastoma cells – electron microscopic tomography reveals 3D arrangement of endoplasmic reticulum, replication organelles, and viral ribonucleoproteins

Abstract text (incl. figure legends and references)

The ultrastructure of two patient-derived glioblastoma cell lines infected with Zika virus (ZIKV) was investigated using scanning transmission electron microscopy (STEM) tomography of high-pressure frozen, freeze-substituted samples. Formation of viral factories containing largely deranged endoplasmic reticulum was observed. In one cell line, newly formed virions and replication organelles (ROs) consisting of luminal vesicles within the ER were found in close proximity to each other. Reconstruction of STEM tomograms showed many about 100-nm-long filaments in the vicinity of ROs, possibly representing viral ribonucleoprotein complexes (RNPs) consisting of the RNA genome and N protein oligomers of ZIKV. The second cell line showed a different morphology upon ZIKV infection, where the virions were found in ER cisternae at a large distance from replication organelles, possibly indicating a dysfunctional transport of virions or different replication kinetics. This study demonstrates the advantage of EM tomography in the investigation of structures like RNPs, which would be hard to identify in conventional two dimensional TEM. STEM tomography in particular is capable of showing the three dimensional arrangement of multiple larger structures, like the ROs, virions and RNPs encountered in viral factories. The lack of image forming lenses in STEM allows for thick specimen of up to 1 μm to be analyzed. In TEM tomgraphy, sample thickness is limited by inelastic scattering of the beam inside the sample, which leads to chromatic aberration in the image forming lenses. Since a STEM does not use image forming lenses, chromatic aberration does not occur in image acquisition.

Reference: Wieland, J., Frey, S., Rupp, U. et al. Zika virus replication in glioblastoma cells: electron microscopic tomography shows 3D arrangement of endoplasmic reticulum, replication organelles, and viral ribonucleoproteins. Histochem Cell Biol 156, 527–538 (2021). https://doi.org/10.1007/s00418-021-02028-2