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.

Each of the eight simian hemorrhagic fever virus minor structural proteins is functionally important

“…The E protein sequences of EAV, PRRSV and LDV contain a conserved N-terminal myristoylation site (Thaa et al, 2009). Inhibition of myristoylation during EAV replication decreased virus infectivity and plaque size but not the amount of extracellular virus suggesting that myristoylation of E is not required for virion budding or release but facilitates infectivity (Thaa et al, 2009). The SHFV E protein sequence contains the conserved N-terminal myristoylation site but the SHFV E′ protein sequence does not.…”

Inhibition of N‐myristoyltransferase1 affects dengue virus replication

“…; Thepparit et al., ) to examine NMT1 gene expression in human dendritic cells at different time‐points postinfection. The results revealed the significant upregulation of NMT1 at 1, 12, and 36 hr, which might represent the time period of DENV infection, and which suggests NMT‐related membrane‐associated viral entry (Abou‐Jaoude, Molina, Maurel, & Sureau, ; Maurer‐Stroh & Eisenhaber, ; Thaa et al., ), transcription and endoplasmic reticulum involvement (Moriya et al., ), and viral exocytosis (Figure ).…”

The Myristate Moiety and Amino Terminus of Vaccinia Virus L1 Constitute a Bipartite Functional Region Needed for Entry

“…virus (12,28,45), Junín virus (88), lymphocytic choriomeningitis virus (62), and equine arteritis virus (75), have evolved to capitalize on the unique properties of myristate for efficient cell entry or membrane fusion. The viral membrane fusion protein(s) of VACV has yet to be identified, although the A17-A27 complex (41) and the EFC constituent H2 (50) have been proposed as candidates.…”