scholarly journals Ecto-ganglioside-sialidase activity of herpes simplex virus-transformed hamster embryo fibroblasts.

1976 ◽  
Vol 70 (3) ◽  
pp. 555-561 ◽  
Author(s):  
C L Schengrund ◽  
A Rosenberg ◽  
M A Repman
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dehydrogenase Activity ◽  
Diazonium Salt ◽  
Intact Cells ◽  
Lactate Dehydrogenase Activity ◽  
Sialidase Activity ◽  
Embryo Fibroblasts ◽  
Simplex Virus

Cellular location of ganglioside-sialidase activity was determined in confluent hamster embryo fibroblasts transformed with herpes simplex virus type 2. Approximately equal specific activities of ganglioside-sialidase activity were found to be associated with the crude lysosomal and crude plasma membrane fractions isolated from whole cell homogenates. Whole transformed cells hydrolyzed exogenous ganglioside substrate, suggesting a partial location of the cellular sialidase on the outer surface of the plasma membrane of these cells. Intact cells were treated with the diazonium salt of sulfanilic acid, a nonpenetrating reagent inhibitory to ecto-enzymes (DePierre, J.W., and M. L. Karnovsky. 1974. J. Biol. Chem. 249:7111-7120). Cytoplasmic lactate dehydrogenase activity was not inhibited by this treatment, and mitochondrial succinate dehydrogenase activity was inhibited only 10%, indicating that intracellular enzymes were not affected. 5'-Nucleotidase activity was diminished 90%, and sialidase very rapidly lost 40% of its exogenously directed activity. These results show that, in herpes simplex virus-transformed fibroblasts, ganglioside-sialidase is both a lysosomal and a plasma membrane enzyme. The plasma membrane sialidase is capable of acting on endogenous plasma membrane sialolipids and also functions in the cultured transformed cell as an ecto-enzyme which can attack exogenous substrates.

scholarly journals Function of Dynein and Dynactin in Herpes Simplex Virus Capsid Transport

2002 ◽  
Vol 13 (8) ◽  
pp. 2795-2809 ◽  
Author(s):  
Katinka Döhner ◽  
André Wolfstein ◽  
Ute Prank ◽  
Christophe Echeverri ◽  
Denis Dujardin ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cytoplasmic Dynein ◽  
Transport Processes ◽  
Viral Envelope ◽  
Cell Periphery ◽  
Virus Binding ◽  
Viral Protein Synthesis ◽  
Simplex Virus

After fusion of the viral envelope with the plasma membrane, herpes simplex virus type 1 (HSV1) capsids are transported along microtubules (MTs) from the cell periphery to the nucleus. The motor ATPase cytoplasmic dynein and its multisubunit cofactor dynactin mediate most transport processes directed toward the minus-ends of MTs. Immunofluorescence microscopy experiments demonstrated that HSV1 capsids colocalized with cytoplasmic dynein and dynactin. We blocked the function of dynein by overexpressing the dynactin subunit dynamitin, which leads to the disruption of the dynactin complex. We then infected such cells with HSV1 and measured the efficiency of particle binding, virus entry, capsid transport to the nucleus, and the expression of immediate-early viral genes. High concentrations of dynamitin and dynamitin-GFP reduced the number of viral capsids transported to the nucleus. Moreover, viral protein synthesis was inhibited, whereas virus binding to the plasma membrane, its internalization, and the organization of the MT network were not affected. We concluded that incoming HSV1 capsids are propelled along MTs by dynein and that dynein and dynactin are required for efficient viral capsid transport to the nucleus.

scholarly journals Herpes simplex virus-1 pUL56 degrades GOPC to alter the plasma membrane proteome

2019 ◽  
Author(s):  
Timothy K. Soh ◽  
Colin T. R. Davies ◽  
Julia Muenzner ◽  
Viv Connor ◽  
Clément R. Bouton ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Host Cell ◽  
Cellular Trafficking ◽  
Toll Like Receptor ◽  
Infected Cells ◽  
Toll Like Receptor 2 ◽  
Simplex Virus ◽  
Hsv 1

SummaryHerpesviruses are ubiquitous in the human population and they extensively remodel the cellular environment during infection. Multiplexed quantitative proteomic analysis over a whole time-course of herpes simplex virus (HSV)-1 infection was used to characterize changes in the host-cell proteome and to probe the kinetics of viral protein production. Several host-cell proteins were targeted for rapid degradation by HSV-1, including the cellular trafficking factor GOPC. We identify that the poorly-characterized HSV-1 protein pUL56 binds directly to GOPC, stimulating its ubiquitination and proteasomal degradation. Plasma membrane profiling revealed that pUL56 mediates specific changes to the surface proteome of infected cells, including loss of IL18 receptor and Toll-like receptor 2, and delivery of Toll-like receptor 2 to the cell-surface requires GOPC. Our study highlights an unanticipated and efficient mechanism whereby a single virus protein targets a cellular trafficking factor to modify the abundance of multiple signaling molecules at the surface of infected cells.

Sialidase activity of oncogenic cells transformed by herpes simplex virus

Virology ◽  
1974 ◽  
Vol 58 (2) ◽  
pp. 595-599 ◽  
Author(s):  
C.-L. Schengrund ◽  
R. Duff ◽  
A. Rosenberg
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Sialidase Activity ◽  
Simplex Virus

scholarly journals Endocytic Internalization of Herpes Simplex Virus 1 in Human Keratinocytes at Low Temperature

2020 ◽  
Author(s):  
Nydia De La Cruz ◽  
Dagmar Knebel-Mörsdorf
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Low Temperature ◽  
Human Keratinocytes ◽  
Herpes Simplex Virus 1 ◽  
Successful Infection ◽  
Simplex Virus ◽  
The Impact ◽  
Hsv 1

Herpes simplex virus 1 (HSV-1) can adopt a variety of pathways to accomplish cellular internalization. In human keratinocytes representing the natural target cell of HSV-1, both direct plasma membrane fusion and endocytic uptake have been found. The impact of either pathway in successful infection, however, remains to be fully understood. To address the role of each internalization mode, we performed infection studies at low temperature as a tool to interfere with endocytic pathways. Interestingly, successful HSV-1 entry in primary human keratinocytes and HaCaT cells was observed even at 7°C, although delayed compared to infection at 37°C. Moreover, ex vivo infection of murine epidermis demonstrated that virus entry at 7°C is not only accomplished in cultured cells but also in tissue. Control experiments with cholera toxin B confirmed a block of endocytic uptake at 7°C. In addition, uptake of dextran by macropinosomes and phagocytic uptake of latex beads was also inhibited at 7°C. Infection of nectin-1-deficient murine keratinocytes affirmed that the entry at 7°C was receptor-dependent. Strikingly, the lysosomotropic agent, ammonium chloride, strongly inhibited HSV-1 entry suggesting a role for endosomal acidification. Ultrastructural analyses in turn revealed free capsids in the cytoplasm as well as virus particles in vesicles after infection at 7°C supporting both plasma membrane fusion and endocytic internalization as already observed at 37°C. Overall, entry of HSV-1 at 7°C suggests that the virus can efficiently adopt nectin-1-dependent unconventional vesicle uptake mechanisms in keratinocytes strengthening the role of endocytic internalization for successful infection. IMPORTANCE The human pathogen herpes simplex virus 1 (HSV-1) relies on multiple internalization pathways to initiate infection. Our focus is on the entry in human keratinocytes, the major in vivo target during primary and recurrent infection. While antivirals reduce the severity of clinical cases, there is no cure or vaccine against HSV. To develop strategies that interfere with virus penetration, we need to understand the various parameters and conditions that determine virus entry. Here, we addressed the impact of virus internalization via vesicles by blocking endocytic processes at low temperature. Intriguingly, we detected entry of HSV-1 even at 7°C which led to infection of primary keratinocytes and epidermal tissue. Moreover, electron microscopy of human keratinocytes at 7°C support that internalization is based on fusion of the viral envelope with the plasma membrane as well as vesicle membranes. These results provide novel insights into conditions that still allow endocytic internalization of HSV-1.

scholarly journals Myosin Va Enhances Secretion of Herpes Simplex Virus 1 Virions and Cell Surface Expression of Viral Glycoproteins

2010 ◽  
Vol 84 (19) ◽  
pp. 9889-9896 ◽  
Author(s):  
Kari L. Roberts ◽  
Joel D. Baines
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Surface Expression ◽  
Dominant Negative ◽  
Cortical Actin ◽  
Binding Domains ◽  
Myosin Va ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT The final step in the egress of herpes simplex virus (HSV) virions requires virion-laden vesicles to bypass cortical actin and fuse with the plasma membrane, releasing virions into the extracellular space. Little is known about the host or viral proteins involved. In the current study, we noted that the conformation of myosin Va (myoVa), a protein known to be involved in melanosome and secretory granule trafficking to the plasma membrane in melanocytes and neuroendocrine cells, respectively, was altered by 4 h after infection with HSV-1 such that an N-terminal epitope expected to be masked in its inactive state was rendered immunoreactive. Wild-type myoVa localized throughout the cytoplasm and to a limited extent in the nuclei of HSV-infected cells. Two different dominant negative myoVa molecules containing cargo-binding domains but lacking the lever arms and actin-binding domains colocalized with markers of the trans-Golgi network (TGN). Expression of dominant negative myoVa isoforms reduced secretion of HSV-1 infectivity into the medium by 50 to 75%, reduced surface expression of glycoproteins B, M, and D, and increased intracellular virus infectivity to levels consistent with increased retention of virions in the cytoplasm. These data suggest that myoVa is activated during HSV-1 infection to help transport virion- and glycoprotein-laden vesicles from the TGN, through the cortical actin, to the plasma membrane. We cannot exclude a role for myoVa in promoting fusion of these vesicles with the inner surface of the plasma membrane. These data also indicate that myoVa is involved in exocytosis in human epithelial cells as well as other cell types.

scholarly journals Herpes Simplex Virus Type 1 Capsids Transit by the trans-Golgi Network, Where Viral Glycoproteins Accumulate Independently of Capsid Egress

2005 ◽  
Vol 79 (14) ◽  
pp. 8847-8860 ◽  
Author(s):  
Sophie Turcotte ◽  
Josée Letellier ◽  
Roger Lippé
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Nuclear Membrane ◽  
Herpes Simplex Virus Type ◽  
Viral Glycoproteins ◽  
Simplex Virus ◽  
Golgi Network

ABSTRACT Egress of herpes capsids from the nucleus to the plasma membrane is a complex multistep transport event that is poorly understood. The current model proposes an initial envelopment at the inner nuclear membrane of capsids newly assembled in the nucleus. The capsids are then released in cytosol by fusion with the outer nuclear membrane. They are finally reenveloped at a downstream organelle before traveling to the plasma membrane for their extracellular release. Although the trans-Golgi network (TGN) is often cited as a potential site of reenvelopment, other organelles have also been proposed, including the Golgi, endoplasmic reticulum-Golgi intermediate compartment, aggresomes, tegusomes, and early or late endosomes. To clarify this important issue, we followed herpes simplex virus type 1 egress by immunofluorescence under conditions that slowed intracellular transport and promoted the accumulation of the otherwise transient reenvelopment intermediate. The data show that the capsids transit by the TGN and point to this compartment as the main reenvelopment site, although a contribution by endosomes cannot formally be excluded. Given that viral glycoproteins are expected to accumulate where capsids acquire their envelope, we examined this prediction and found that all tested could indeed be detected at the TGN. Moreover, this accumulation occurred independently of capsid egress. Surprisingly, capsids were often found immediately adjacent to the viral glycoproteins at the TGN.

scholarly journals Intracellular Trafficking of the UL11 Tegument Protein of Herpes Simplex Virus Type 1

2001 ◽  
Vol 75 (24) ◽  
pp. 12209-12219 ◽  
Author(s):  
Joshua S. Loomis ◽  
J. Bradford Bowzard ◽  
Richard J. Courtney ◽  
John W. Wills
Keyword(s):  
Plasma Membrane ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Golgi Apparatus ◽  
Membrane Trafficking ◽  
Intracellular Trafficking ◽  
Tegument Protein ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Growing evidence indicates that herpes simplex virus type 1 (HSV-1) acquires its final envelope in the trans-Golgi network (TGN). During the envelopment process, the viral nucleocapsid as well as the envelope and tegument proteins must arrive at this site in order to be incorporated into assembling virions. To gain a better understanding of how these proteins associate with cellular membranes and target to the correct compartment, we have been studying the intracellular trafficking properties of the small tegument protein encoded by the UL11 gene of HSV-1. This 96-amino-acid, myristylated protein accumulates on the cytoplasmic face of internal membranes, where it is thought to play a role in nucleocapsid envelopment and egress. When expressed in the absence of other HSV-1 proteins, the UL11 protein localizes to the Golgi apparatus, and previous deletion analyses have revealed that the membrane-trafficking information is contained within the first 49 amino acids. The goal of this study was to map the functional domains required for proper Golgi membrane localization. In addition to N-terminal myristylation, which allows for weak membrane binding, UL11 appears to be palmitylated on one or more of three consecutive N-terminal cysteines. Using membrane-pelleting experiments and confocal microscopy, we show that palmitylation of UL11 is required for both Golgi targeting specificity and strong membrane binding. Furthermore, we found that a conserved acidic cluster within the first half of UL11 is required for the recycling of this tegument protein from the plasma membrane to the Golgi apparatus. Taken together, our results demonstrate that UL11 has highly dynamic membrane-trafficking properties, which suggests that it may play multiple roles on the plasma membrane as well as on the nuclear and TGN membranes.

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