The coronavirus membrane protein (M) is the key player in the assembly of virions at intracellular membranes between endoplasmic-reticulum and Golgi-complex. In this study, the topology, processing and intracellular transport of SARS-Coronavirus M was characterized. It could be shown that M consists of a short N-terminal ectodomain with a single N-glycosylation site at position four, three transmembrane domains and a C-terminal endodomain. After recombinant expression and in SARS-CoV infected cells, M exhibited an unmodified and a N-glycosylated form and accumulated at steady state in Golgi apparatus. Further analysis revealed that recombinant M is not completely retained in the Golgi-complex but further transported to the plasma membrane. By means of two different antibody uptake approaches, M was detected intracellularly suggesting efficient endocytosis from the cell surface. In addition, immunofluorescence analysis demonstrated that M was able to retain the viral spike-protein in perinuclear regions where budding of new virions takes place.
Using an infectious cDNA clone of SARS-CoV (constructed in the lab of Prof. C. Drosten, Bernhard-Nocht Institute Hamburg), a recombinant virus with a glycosylation-deficient M was rescued. Electron microscopy analysis, plaque assays and growth kinetics do not show pivotal differences in the morphology and the replication of this mutant virus in comparison to wild type recombinant SARS-CoV.
Voß, Daniel: Glykosylierung und intrazellulärer Transport des SARS-Coronavirus Membranproteins. : Philipps-Universität Marburg 2008-01-10. DOI: https://doi.org/10.17192/z2007.0782.
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