title: Myristoylation of the arterivirus E protein: the fatty acid modification is not essential for membrane association but contributes significantly to virus infectivity.
abstract: The envelope of equine arteritis virus (EAV) contains two glycoprotein complexes (GP2b/GP3/GP4 and GP5/M) and the small, non-glycosylated E protein. As E is essential for the production of infectious progeny but dispensable for assembly and release of virus-like particles, it probably mediates virus entry into cells, putatively in concert with the GP2b/GP3/GP4 complex. The E protein contains a central hydrophobic domain and a conserved potential site for N-terminal myristoylation, a hydrophobic modification usually pivotal for membrane targeting of the modified protein. Here, it was shown by radiolabelling that E is myristoylated at glycine-2, both in transfected cells as a fusion protein with yellow fluorescent protein (YFP) and in virus particles. Biochemical fractionation revealed that E-YFP with an inactivated acylation site was still completely membrane-bound, indicating that the putative transmembrane domain of E mediates membrane targeting. Confocal microscopy showed that both myristoylated and non-myristoylated E-YFP were localized to the endoplasmic reticulum and Golgi complex, the membranes from which EAV buds. The presence of a myristoylation inhibitor during replication of EAV, whilst completely blocking E acylation, reduced virus titres by 1.5 log(10). Similarly, a mutant EAV with non-myristoylatable E grew to a titre five- to sevenfold lower than that of the wild-type virus and exhibited a reduced plaque size. Western blotting of cell-culture supernatants showed that N and M, the major structural proteins of EAV, are released in similar amounts by cells transfected with wild-type and mutant genomes. Thus, E myristoylation is not required for budding of particles and probably has a function during virus entry.
authors: Department of Immunology and Molecular Biology Veterinary Faculty, Free University, Berlin, Germany.
authors: Bastian
authors: B
authors: Thaa
authors: Department of Immunology and Molecular Biology Veterinary Faculty, Free University, Berlin, Germany.
authors: Aleksander
authors: A
authors: Kabatek
authors: Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, The Netherlands.
authors: Jessika C
authors: JC
authors: Zevenhoven-Dobbe
authors: Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, The Netherlands.
authors: Eric J
authors: EJ
authors: Snijder
authors: Department of Biology, Molecular Biophysics, Humboldt University, Berlin, Germany.
authors: Andreas
authors: A
authors: Herrmann
authors: Department of Immunology and Molecular Biology Veterinary Faculty, Free University, Berlin, Germany.
authors: Michael
authors: M
authors: Veit
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004721:Endoplasmic Reticulum / Q000821:virology; D007240:Equartevirus / Q000472:pathogenicity*; D005227:Fatty Acids / Q000378:metabolism*; D006056:Golgi Apparatus / Q000821:virology; D007425:Intracellular Membranes / Q000821:virology; D011499:Protein Processing, Post-Translational; D015678:Viral Structural Proteins / Q000378:metabolism*; D053586:Virus Internalization
chemical_list: C120679:E protein, Equine arteritis virus; D005227:Fatty Acids; D015678:Viral Structural Proteins