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Nelfinavir impairs glycosylation of herpes simplex virus 1 envelope proteins and blocks virus maturation.

Gantt S, Gachelet E, Carlsson J, Barcy S, Casper C, Lagunoff M - Adv Virol (2015)

Bottom Line: NFV did not significantly affect the level of expression of late HSV-1 gene products.NFV was found to decrease glycosylation of viral glycoproteins B and C and resulted in aberrant subcellular localization, consistent with induction of endoplasmic reticulum stress and the unfolded protein response by NFV.These results demonstrate that NFV causes alterations in HSV-1 glycoprotein maturation and egress and likely acts on one or more host cell functions that are important for HHV replication.

View Article: PubMed Central - PubMed

Affiliation: Seattle Children's Research Institute, University of Washington, Seattle, WA 98101, USA ; Department of Pediatrics, University of Washington, Seattle, WA 98105, USA ; Department of Global Health, University of Washington, Seattle, WA 98195, USA.

ABSTRACT
Nelfinavir (NFV) is an HIV-1 aspartyl protease inhibitor that has numerous effects on human cells, which impart attractive antitumor properties. NFV has also been shown to have in vitro inhibitory activity against human herpesviruses (HHVs). Given the apparent absence of an aspartyl protease encoded by HHVs, we investigated the mechanism of action of NFV herpes simplex virus type 1 (HSV-1) in cultured cells. Selection of HSV-1 resistance to NFV was not achieved despite multiple passages under drug pressure. NFV did not significantly affect the level of expression of late HSV-1 gene products. Normal numbers of viral particles appeared to be produced in NFV-treated cells by electron microscopy but remain within the cytoplasm more often than controls. NFV did not inhibit the activity of the HSV-1 serine protease nor could its antiviral activity be attributed to inhibition of Akt phosphorylation. NFV was found to decrease glycosylation of viral glycoproteins B and C and resulted in aberrant subcellular localization, consistent with induction of endoplasmic reticulum stress and the unfolded protein response by NFV. These results demonstrate that NFV causes alterations in HSV-1 glycoprotein maturation and egress and likely acts on one or more host cell functions that are important for HHV replication.

No MeSH data available.


Related in: MedlinePlus

NFV treatment impairs HSV-1 maturation and egress. Shown are representative transmission election micrographs, performed 20 hours after HSV-1 infection of HF that were either untreated (panels (a) and (c)) or treated with 10 μM NFV (panels (b) and (d)). Low power micrographs are shown in panels (a) and (b); bar = 2 μm. White arrows indicate HSV-1 capsids in the nucleus (N), black arrowheads indicate capsids in the cytoplasm (C), and black arrows indicate extracellular (E) virions. Similar numbers of capsids were observed in the nucleus of untreated and NFV-treated cells (approximately 39 and 44, resp.). However, untreated cells were observed to have fewer capsids in the cytoplasm (~14 versus 79) and substantially more extracellular virus particles (~51 versus 3). Cytoplasmic virus particles are shown in panels (c) and (d); bar = 0.5 μm. Enveloped virus particles (indicated in panel (c) by a black arrow) were commonly observed in untreated cells but rarely in NFV-treated cells (approximately 9 versus 0, resp., in the fields shown). Capsids in the cytoplasm of NFV-treated cells were almost exclusively nonenveloped (indicated by arrowheads; approximately 16 versus 23 in panels (c) and (d), resp.).
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fig4: NFV treatment impairs HSV-1 maturation and egress. Shown are representative transmission election micrographs, performed 20 hours after HSV-1 infection of HF that were either untreated (panels (a) and (c)) or treated with 10 μM NFV (panels (b) and (d)). Low power micrographs are shown in panels (a) and (b); bar = 2 μm. White arrows indicate HSV-1 capsids in the nucleus (N), black arrowheads indicate capsids in the cytoplasm (C), and black arrows indicate extracellular (E) virions. Similar numbers of capsids were observed in the nucleus of untreated and NFV-treated cells (approximately 39 and 44, resp.). However, untreated cells were observed to have fewer capsids in the cytoplasm (~14 versus 79) and substantially more extracellular virus particles (~51 versus 3). Cytoplasmic virus particles are shown in panels (c) and (d); bar = 0.5 μm. Enveloped virus particles (indicated in panel (c) by a black arrow) were commonly observed in untreated cells but rarely in NFV-treated cells (approximately 9 versus 0, resp., in the fields shown). Capsids in the cytoplasm of NFV-treated cells were almost exclusively nonenveloped (indicated by arrowheads; approximately 16 versus 23 in panels (c) and (d), resp.).

Mentions: As shown by Kalu et al. [25] and supported by our preliminary experiments (data not shown), late HSV-1 gene expression in HF cells was not reduced by NFV treatment. We therefore explored the effects of NFV on HSV-1 infected HF cells by transmission electron microscopy. Normal numbers of virus particles appeared to be produced in NFV-treated cells. Similar numbers of capsids were observed in the nucleus of untreated and NFV-treated cells (approximately 39 and 44, resp., Figures 4(a) and 4(b)). However, compared with untreated cells, capsids in NFV-treated cells appeared to be disproportionately retained within the cytoplasm (~14 versus 79, resp.), in which virus particles were more often located outside the plasma membrane (~51 versus 3), suggesting a block in virus maturation or egress. In addition, in contrast to those in the cytoplasm of untreated cells, cytoplasmic capsids in NFV-treated cells were rarely observed to be enveloped (~9 versus 0 in the fields shown; Figures 4(c) and 4(d)). Interestingly, although endoplasmic reticulum (ER) stress, the unfolded protein response (UPR), and autophagy are well known effects of NFV [16, 26–29], neither ER dilation nor the abundance of double-membrane bound vesicles consistent with autophagosomes appeared consistently different between NVF-treated and untreated HSV-1-infected cells.


Nelfinavir impairs glycosylation of herpes simplex virus 1 envelope proteins and blocks virus maturation.

Gantt S, Gachelet E, Carlsson J, Barcy S, Casper C, Lagunoff M - Adv Virol (2015)

NFV treatment impairs HSV-1 maturation and egress. Shown are representative transmission election micrographs, performed 20 hours after HSV-1 infection of HF that were either untreated (panels (a) and (c)) or treated with 10 μM NFV (panels (b) and (d)). Low power micrographs are shown in panels (a) and (b); bar = 2 μm. White arrows indicate HSV-1 capsids in the nucleus (N), black arrowheads indicate capsids in the cytoplasm (C), and black arrows indicate extracellular (E) virions. Similar numbers of capsids were observed in the nucleus of untreated and NFV-treated cells (approximately 39 and 44, resp.). However, untreated cells were observed to have fewer capsids in the cytoplasm (~14 versus 79) and substantially more extracellular virus particles (~51 versus 3). Cytoplasmic virus particles are shown in panels (c) and (d); bar = 0.5 μm. Enveloped virus particles (indicated in panel (c) by a black arrow) were commonly observed in untreated cells but rarely in NFV-treated cells (approximately 9 versus 0, resp., in the fields shown). Capsids in the cytoplasm of NFV-treated cells were almost exclusively nonenveloped (indicated by arrowheads; approximately 16 versus 23 in panels (c) and (d), resp.).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig4: NFV treatment impairs HSV-1 maturation and egress. Shown are representative transmission election micrographs, performed 20 hours after HSV-1 infection of HF that were either untreated (panels (a) and (c)) or treated with 10 μM NFV (panels (b) and (d)). Low power micrographs are shown in panels (a) and (b); bar = 2 μm. White arrows indicate HSV-1 capsids in the nucleus (N), black arrowheads indicate capsids in the cytoplasm (C), and black arrows indicate extracellular (E) virions. Similar numbers of capsids were observed in the nucleus of untreated and NFV-treated cells (approximately 39 and 44, resp.). However, untreated cells were observed to have fewer capsids in the cytoplasm (~14 versus 79) and substantially more extracellular virus particles (~51 versus 3). Cytoplasmic virus particles are shown in panels (c) and (d); bar = 0.5 μm. Enveloped virus particles (indicated in panel (c) by a black arrow) were commonly observed in untreated cells but rarely in NFV-treated cells (approximately 9 versus 0, resp., in the fields shown). Capsids in the cytoplasm of NFV-treated cells were almost exclusively nonenveloped (indicated by arrowheads; approximately 16 versus 23 in panels (c) and (d), resp.).
Mentions: As shown by Kalu et al. [25] and supported by our preliminary experiments (data not shown), late HSV-1 gene expression in HF cells was not reduced by NFV treatment. We therefore explored the effects of NFV on HSV-1 infected HF cells by transmission electron microscopy. Normal numbers of virus particles appeared to be produced in NFV-treated cells. Similar numbers of capsids were observed in the nucleus of untreated and NFV-treated cells (approximately 39 and 44, resp., Figures 4(a) and 4(b)). However, compared with untreated cells, capsids in NFV-treated cells appeared to be disproportionately retained within the cytoplasm (~14 versus 79, resp.), in which virus particles were more often located outside the plasma membrane (~51 versus 3), suggesting a block in virus maturation or egress. In addition, in contrast to those in the cytoplasm of untreated cells, cytoplasmic capsids in NFV-treated cells were rarely observed to be enveloped (~9 versus 0 in the fields shown; Figures 4(c) and 4(d)). Interestingly, although endoplasmic reticulum (ER) stress, the unfolded protein response (UPR), and autophagy are well known effects of NFV [16, 26–29], neither ER dilation nor the abundance of double-membrane bound vesicles consistent with autophagosomes appeared consistently different between NVF-treated and untreated HSV-1-infected cells.

Bottom Line: NFV did not significantly affect the level of expression of late HSV-1 gene products.NFV was found to decrease glycosylation of viral glycoproteins B and C and resulted in aberrant subcellular localization, consistent with induction of endoplasmic reticulum stress and the unfolded protein response by NFV.These results demonstrate that NFV causes alterations in HSV-1 glycoprotein maturation and egress and likely acts on one or more host cell functions that are important for HHV replication.

View Article: PubMed Central - PubMed

Affiliation: Seattle Children's Research Institute, University of Washington, Seattle, WA 98101, USA ; Department of Pediatrics, University of Washington, Seattle, WA 98105, USA ; Department of Global Health, University of Washington, Seattle, WA 98195, USA.

ABSTRACT
Nelfinavir (NFV) is an HIV-1 aspartyl protease inhibitor that has numerous effects on human cells, which impart attractive antitumor properties. NFV has also been shown to have in vitro inhibitory activity against human herpesviruses (HHVs). Given the apparent absence of an aspartyl protease encoded by HHVs, we investigated the mechanism of action of NFV herpes simplex virus type 1 (HSV-1) in cultured cells. Selection of HSV-1 resistance to NFV was not achieved despite multiple passages under drug pressure. NFV did not significantly affect the level of expression of late HSV-1 gene products. Normal numbers of viral particles appeared to be produced in NFV-treated cells by electron microscopy but remain within the cytoplasm more often than controls. NFV did not inhibit the activity of the HSV-1 serine protease nor could its antiviral activity be attributed to inhibition of Akt phosphorylation. NFV was found to decrease glycosylation of viral glycoproteins B and C and resulted in aberrant subcellular localization, consistent with induction of endoplasmic reticulum stress and the unfolded protein response by NFV. These results demonstrate that NFV causes alterations in HSV-1 glycoprotein maturation and egress and likely acts on one or more host cell functions that are important for HHV replication.

No MeSH data available.


Related in: MedlinePlus