Differential maturation and subcellular localization of severe acute respiratory syndrome coronavirus surface proteins S, M and E

Abstract

Post-translational modifications and correct subcellular localization of viral structural proteins are prerequisites for assembly and budding of enveloped viruses. Coronaviruses, like the severe acute respiratory syndrome-associated virus (SARS-CoV), bud from the endoplasmic reticulum-Golgi intermediate compartment. In this study, the subcellular distribution and maturation of SARS-CoV surface proteins S, M and E were analysed by using C-terminally tagged proteins. As early as 30 min post-entry into the endoplasmic reticulum, high-mannosylated S assembles into trimers prior to acquisition of complex N-glycans in the Golgi. Like S, M acquires high-mannose N-glycans that are subsequently modified into complex N-glycans in the Golgi. The N-glycosylation profile and the absence of O-glycosylation on M protein relate SARS-CoV to the previously described group 1 and 3 coronaviruses. Immunofluorescence analysis shows that S is detected in several compartments along the secretory pathway from the endoplasmic reticulum to the plasma membrane while M predominantly localizes in the Golgi, where it accumulates, and in trafficking vesicles. The E protein is not glycosylated. Pulse-chase labelling and confocal microscopy in the presence of protein translation inhibitor cycloheximide revealed that the E protein has a short half-life of 30 min. E protein is found in bright perinuclear patches colocalizing with endoplasmic reticulum markers. In conclusion, SARS-CoV surface proteins S, M and E show differential subcellular localizations when expressed alone suggesting that additional cellular or viral factors might be required for coordinated trafficking to the virus assembly site in the endoplasmic reticulum-Golgi intermediate compartment.

Expression and membrane integration of SARS-CoV E protein and its interaction with M protein

“…Differential maturation and subcellular localization of SARS-CoV S, M, and E proteins is also reported [11]. Both SARS-CoV M and E proteins are integral membrane proteins, though neither of them contains a conventional signal peptide [10,11].…”

“…Both SARS-CoV M and E proteins are integral membrane proteins, though neither of them contains a conventional signal peptide [10,11]. Expression and membrane integration of SARS-CoV M protein was characterized previously [12].…”

The SARS Coronavirus E Protein Interacts with PALS1 and Alters Tight Junction Formation and Epithelial Morphogenesis

“…E is a small hydrophobic integral membrane protein of 76 amino acid residues, which plays a major but not fully understood role in virus morphogenesis and budding (Liu et al , 2007; Siu et al , 2008). In transfected and infected cells, it is known that E localizes at the membranes of the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) and Golgi apparatus, where virus assembly occurs (Nal et al , 2005; Liu et al , 2007; Siu et al , 2008). In this report we study the biochemical interaction between SARS-CoV E and human PALS1 and investigate the consequences of E expression in MDCKII epithelial cells on cyst morphogenesis and tight junction formation.…”

Subcellular Localization of SARS-CoV Structural Proteins

“…1 Our observation of M in the ERGIC agrees with a recent report which also noted an overlap of SARS-CoV M with the ERGIC in addition to localization in the Golgi. 12 SARS-CoV E also localized to the Golgi (Fig. 1, upper right).…”