Brefeldin A affects West Nile virus replication in Vero cells but not C6/36 cells

The intracellular assembly site for flaviviruses in currently not known but is presumed to be located within the lumen of the rough endoplasmic reticulum (RER). Building on previous studies involving immunofluorescence (IF) and cryoimmunoelectron microscopy of Kunjin virus (KUN)-infected cells, we sought to identify the steps involved in the assembly and maturation of KUN. Thus, using antibodies directed against envelope protein E in IF analysis, we found the accumulation of E within regions coincident with the RER and endosomal compartments. Immunogold labeling of cryosections of infected cells indicated that E and minor envelope protein prM were localized to reticulum membranes continuous with KUN-induced convoluted membranes (CM) or paracrystalline arrays (PC) and that sometimes the RER contained immunogold-labeled virus particles. Both proteins were also observed to be labeled in membranes at the periphery of the induced CM or PC structures, but the latter were very seldom labeled internally. Utilizing drugs that inhibit protein and/or membrane traffic throughout the cell, we found that the secretion of KUN particles late in infection was significantly affected in the presence of brefeldin A and that the infectivity of secreted particles was severely affected in the presence of monensin and N-nonyl-deoxynojirimycin. Nocodazole did not appear to affect maturation, suggesting that microtubules play no role in assembly or maturation processes. Subsequently, we showed that the exit of intact virions from the RER involves the transport of individual virions within individual vesicles en route to the Golgi apparatus. The results suggest that the assembly of virions occurs within the lumen of the RER and that subsequent maturation occurs via the secretory pathway.

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“…Sreenivasan et al (51) added BFA at 1 h p.i. to investigate its effects on West Nile virus strain Sarafend replication in Vero cells.…”

Section: Discussionmentioning

confidence: 99%

Assembly and Maturation of the Flavivirus Kunjin Virus Appear To Occur in the Rough Endoplasmic Reticulum and along the Secretory Pathway, Respectively

Mackenzie

Westaway

2001

J Virol

274

236

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“…However, when a study was conducted on the effects of Brefeldin A (BFA) on West Nile (Sarafend) virus replication [18], maturation at the plasma membrane was observed under the transmission electron microscopy. The budding process was inhibited by Brefeldin A (BFA) only in infected Vero cells and not the C6/36 cells.…”

Section: Introductionmentioning

confidence: 99%

Flavivirus West Nile (Sarafend) egress at the plasma membrane

Howe

Sreenivasan

et al. 1994

Archives of Virology

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West Nile (Sarafend) virus was distinctly observed to bud from the plasma membrane rather than mature intracellularly. This has been observed with transmission electron microscopy. Using conventional scanning electron microscopy, budding at the plasma membrane especially at the filopodia was clearly illustrated. Immunogold labelling against the virus envelope protein was also performed to confirm this mode of exit. The gold particles were observed to be located at the sites where virus budding was seen under the field emission scanning electron microscope.

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“…Consistent with results observed in WNV and Kunjin viruses, the treatment of cells with BFA, at even the lowest concentrations tested, resulted in total ablation of infectious virus release ( Figure 5-8) (Hunsperger and Roehrig, 2005;Mackenzie and Westaway, 2001;Sreenivasan et al, 1993). Normally, the virus would acquire its envelope by budding through the lumen of the ER, then moving through the Golgi complex before release.…”

Section: Discussionsupporting

confidence: 68%

“…While the precise site of hexamer formation remains elusive, it is presumed to occur before the trans-Golgi, where the high mannose CHO attached to Asn 130 is trimmed and processed to a complex form. As NS1 moves through the secretory pathway, we hypothesize that membrane associated NS1 concentrates with cholesterol, cells, also found that C6/36 cells were resistant to the effects of BFA (Sreenivasan et al, 1993).…”

Section: Discussionmentioning

confidence: 99%

Dengue virus non-structural protein NS1: Structure, intracellular trafficking and role as a diagnostic marker

Muller¹

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Brefeldin A affects West Nile virus replication in Vero cells but not C6/36 cells

Cited by 17 publications

References 21 publications

Assembly and Maturation of the Flavivirus Kunjin Virus Appear To Occur in the Rough Endoplasmic Reticulum and along the Secretory Pathway, Respectively

Assembly and Maturation of the Flavivirus Kunjin Virus Appear To Occur in the Rough Endoplasmic Reticulum and along the Secretory Pathway, Respectively

Flavivirus West Nile (Sarafend) egress at the plasma membrane

Dengue virus non-structural protein NS1: Structure, intracellular trafficking and role as a diagnostic marker

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