In vitro characterization of the HSV-1 UL53 gene product

Ramaswamy, R S; Cholland, Thomas C.

doi:10.1016/0042-6822(92)90024-j

Cited by 47 publications

(71 citation statements)

References 26 publications

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“…UL53 encodes a glycosylated polypeptide, gK, thought to be essential for virus replication (Hutchinson et al, 1992;Ramaswamy & Holland, 1992). A single amino acid substitution within gK results in extensive cell-to-cell fusion (Debroy et al, 1985;Pogue-Geile & Spear, 1987), suggesting a role in virus penetration, an event involving fusion between the virus and cell membranes.…”

Section: Introductionmentioning

confidence: 99%

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Characterization of the UL10 gene product of herpes simplex virus type 1 and investigation of its role in vivo

MacLean

Robertson

Jamieson

1993

Journal of General Virology

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(ULlO-del). The in vitro growth properties of this virus were consistent with previous studies; it grew to give slightly lower yields than wild-type and revertant viruses, and had no apparent temperaturesensitive or host range phenotype. In vivo, in a mouse model, ULlO-del was capable of establishing a latent infection, although it was impaired for growth at the periphery, and for spread to and/or growth within the nervous system relative to wild-type or revertant viruses.

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Section: Introductionmentioning

confidence: 99%

“…which is probably cleaved (Ramaswamy & Holland, 1992). UL53 encodes a glycosylated polypeptide, gK, thought to be essential for virus replication (Hutchinson et al, 1992;Ramaswamy & Holland, 1992).…”

Section: Introductionmentioning

confidence: 99%

Characterization of the UL10 gene product of herpes simplex virus type 1 and investigation of its role in vivo

MacLean

Robertson

Jamieson

1993

Journal of General Virology

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“…It has characteristics of a glycosylated membrane protein, including a cleavable 30-amino-acid NH 2 -terminal signal sequence, two potential sites for N glycosylation, and several hydrophobic domains (hpd) (8,36). Initially, gK was predicted to have four transmembrane-spanning regions; however, experiments with in vitro-translated gK in the presence of microsomal membranes suggested that gK contained three instead of four membrane-spanning regions (8,32,37). Cleavage of the NH 2 -terminal signal peptide and the addition of carbohydrates at amino acid residues 48 and 58 suggested that the amino-terminal domain (domain I) is an ectodomain (10,18,37).…”

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confidence: 99%

“…Initially, gK was predicted to have four transmembrane-spanning regions; however, experiments with in vitro-translated gK in the presence of microsomal membranes suggested that gK contained three instead of four membrane-spanning regions (8,32,37). Cleavage of the NH 2 -terminal signal peptide and the addition of carbohydrates at amino acid residues 48 and 58 suggested that the amino-terminal domain (domain I) is an ectodomain (10,18,37). The presence of multiple syncytial mutations within gK implied that gK is expressed on infected-cell surfaces where it could directly be involved in virus-induced cell fusion.…”

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confidence: 99%

Plasma Membrane Topology of Syncytial Domains of Herpes Simplex Virus Type 1 Glycoprotein K (gK): the UL20 Protein Enables Cell Surface Localization of gK but Not gK-Mediated Cell-to-Cell Fusion

Foster

Álvarez

Kousoulas

2003

J Virol

117

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Most spontaneously occurring mutations that cause extensive herpes simplex virus type 1 (HSV-1)-induced cell fusion are single amino acid changes within glycoprotein K (gK). Despite the strong genetic association of gK with virus-induced cell fusion, its direct involvement in cellular membrane fusion has been controversial, largely due to previously unsuccessful efforts to detect gK expression on virion and cellular surfaces. Recently, we showed that gK is expressed on HSV-1 virions and functioned in virus entry (T. P. Foster, G. V. Rybachuk, and K. G. Kousoulas, J. Virol. 75:12431-12438, 2001). To determine whether gK is expressed on cellular surfaces, as well as its membrane topology, we generated the recombinant viruses gKV5DI, gKV5DII, gKV5DIII, and gKV5DIVcontaining insertions of the V5 antigenic epitope within each of four domains of gK predicted to localize either in the cytoplasmic side or in the extracytoplasmic side of cellular membranes. Immunohistochemical and confocal microscopy analyses of infected cells showed that both wild-type and syncytial forms of gK were expressed on cell surfaces. Analysis of the topology of the V5-tagged gK revealed that gK domains I and IV were located extracellularly, whereas domains II and III were localized intracellularly. Transiently expressed gK failed to localize in cellular plasma membranes. In contrast, infection of gKtransfected cells with the gK-null virus ⌬gK enabled expression of gK on cell surfaces, as well as gK-mediated membrane fusion. Transient-coexpression experiments revealed that the UL20 protein enabled cell surface expression of gK, but not gK-mediated cell-to-cell fusion, indicating that additional viral proteins are required for expression of the gK syncytial phenotype.Viral glycoproteins are key determinants of membrane fusion events throughout the life cycle of herpesviruses. Herpes simplex viruses (HSVs) specify at least 11 glycoproteins:

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“…Mutations that cause extensive virus-induced cell-to-cell fusion have been mapped to at least four regions of the viral genome: the UL20 gene (1,50,53), the UL24 gene (38, 64), the UL27 gene encoding glycoprotein B (gB) (9, 57), and the UL53 gene coding for gK (3,16,34,58,59,63). Of these four membraneassociated proteins, UL20p and gK are essential for the intracellular transport of virions to extracellular spaces (1,25,29,35,39,55).The UL20 and UL53(gK) genes encode multipass transmembrane proteins of 222 and 338 amino acids, respectively, that are conserved in all alphaherpesviruses (16,50,60). Although both proteins have multiple sites where posttranslational modification can occur, only gK is posttranslationally modified by N-linked carbohydrate addition (16,34,60).…”

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Herpes Simplex Virus Type 1 Glycoprotein K and the UL20 Protein Are Interdependent for Intracellular Trafficking andtrans-Golgi Network Localization

Foster

Melancon

Olivier

et al. 2004

J Virol

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Final envelopment of the cytoplasmic herpes simplex virus type 1 (HSV-1) nucleocapsid is thought to occur by budding into trans-Golgi network (TGN)-derived membranes. The highly membrane-associated proteins UL20p and glycoprotein K (gK) are required for cytoplasmic envelopment at the TGN and virion transport from the TGN to extracellular spaces. Furthermore, the UL20 protein is required for intracellular transport and cell surface expression of gK. Independently expressed gK or UL20p via transient expression in Vero cells failed to be transported from the endoplasmic reticulum (ER). Similarly, infection of Vero cells with either gK-null or UL20-null viruses resulted in ER entrapment of UL20p or gK, respectively. In HSV-1 wild-type virus infections and to a lesser extent in transient gK and UL20p coexpression experiments, both gK and UL20p localized to the Golgi apparatus. In wild-type, but not UL20-null, viral infections, gK was readily detected on cell surfaces. In contrast, transiently coexpressed gK and UL20p predominantly localized to the TGN and were not readily detected on cell surfaces. However, TGN-localized gK and UL20p originated from endocytosed gK and UL20p expressed at cell surfaces. Retention of UL20p to the ER through the addition of an ER retention motif forced total ER retention of gK, indicating that transport of gK is absolutely dependent on UL20p transport. In all experiments, gK and UL20p colocalized at intracellular sites, including the ER, Golgi, and TGN. These results are consistent with the hypothesis that gK and UL20p directly interact and that this interaction facilitates their TGN localization, an important prerequisite for cytoplasmic virion envelopment and egress.It is widely accepted that herpesvirus egress entails a twostage envelopment process. Initially, capsids assemble within the nuclei and virions acquire an initial envelope by budding into the perinuclear spaces. Subsequently, these enveloped virions lose their envelope by fusion with the outer nuclear lamellae. Within the cytoplasm, tegument proteins associate with the viral nucleocapsid and final envelopment occurs by budding of cytoplasmic capsids into specific trans-Golgi network (TGN)-associated membranes. Mature virions subsequently traffic to cell surfaces, presumably following the cellular secretory pathway (31,54,65). Although several lines of evidence support this model of TGN assembly and egress (8,30,54,76), the specific viral and cellular mechanisms responsible for the targeted trafficking of viral envelope glycoproteins to intracellular sites of virion envelopment are only recently being elucidated.Herpes simplex viruses (HSVs) specify at least 11 virally encoded glycoproteins, as well as several nonglycosylated membrane-associated proteins, that are pivotal in membrane fusion processes during a productive viral infection. Mutations that cause extensive virus-induced cell-to-cell fusion have been mapped to at least four regions of the viral genome: the UL20 gene (1,50,53), the UL24 gene (38, 64), the UL27 gene ...

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In vitro characterization of the HSV-1 UL53 gene product

Cited by 47 publications

References 26 publications

Characterization of the UL10 gene product of herpes simplex virus type 1 and investigation of its role in vivo

Characterization of the UL10 gene product of herpes simplex virus type 1 and investigation of its role in vivo

Plasma Membrane Topology of Syncytial Domains of Herpes Simplex Virus Type 1 Glycoprotein K (gK): the UL20 Protein Enables Cell Surface Localization of gK but Not gK-Mediated Cell-to-Cell Fusion

Herpes Simplex Virus Type 1 Glycoprotein K and the UL20 Protein Are Interdependent for Intracellular Trafficking andtrans-Golgi Network Localization

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