Human Cytomegalovirus: Purification of Enveloped Virions and Dense Bodies

Talbot, Prue; Almeida, June D.

doi:10.1099/0022-1317-36-2-345

Cited by 129 publications

(121 citation statements)

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“…Our analysis has not been extended to include betaand gammaherpesviruses, although CMV-infected cells do produce particles (termed dense bodies) which lack capsids (Talbot & Almeida, 1977;Irmiere & Gibson, 1983). Dense bodies are considerably larger than CMV virions and have a simpler composition, consisting predominantly of an Mr 69K matrix protein (Irmiere & Gibson, 1983).…”

Section: Discussionmentioning

confidence: 99%

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Characterization of enveloped tegument structures (L particles) produced by alphaherpesviruses: integrity of the tegument does not depend on the presence of capsid or envelope

McLauchlan

Rixon

1992

Journal of General Virology

108

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Recent studies have shown that infection with herpes simplex virus type 1 (HSV-1) strain 17 generates in addition to virions a novel type of non-infectious particle. These particles, termed L particles, lack capsids and viral DNA, and consist predominantly of tegument and envelope proteins. We show that L particle production is not restricted to one strain of HSV-1, and that pseudorabies virus and equine herpesvirus type 1 also release particles which are similar in composition to and morphologically indistinguishable from HSV-1 L particles. Data obtained from monoclonal antibody analysis revealed that Vmw175, an immediate early HSV-1 polypeptide which had been previously identified as a virion component, is located predominantly in L particles and not in virions. Following removal of the envelope from L particles, the remaining tegument material largely retained its structural integrity, indicating that the structure of the tegument does not depend on the presence of the capsid or envelope.

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Section: Discussionmentioning

confidence: 99%

“…In CMV infections two types of enveloped particle have been identified in addition to mature virions (Talbot & Almeida, 1977;Irmiere & Gibson, 1983). These have been termed dense bodies and noninfectious enveloped particles (NIEPs).…”

Section: Introductionmentioning

confidence: 99%

Characterization of enveloped tegument structures (L particles) produced by alphaherpesviruses: integrity of the tegument does not depend on the presence of capsid or envelope

McLauchlan

Rixon

1992

Journal of General Virology

108

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“…The mature viruses were purified by a modified rate-velocity glyceroltartrate gradient centrifugation as described previously (Irmiere & Gibson, 1983;Talbot & Almeida, 1977;Wang et al, 2004). Virion particles were subjected to a second gradient centrifugation in order to ensure their purity.…”

Section: Methodsmentioning

confidence: 99%

Novel virus-associated proteins encoded by UL112–113 of human cytomegalovirus

Wang

et al. 2009

Journal of General Virology

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Evidence suggests that the products of the human cytomegalovirus (HCMV) UL112-113 genes are involved in viral DNA replication during lytic infection. A polyclonal antibody was raised against the UL112 open reading frame (ORF) to characterize its function in detail. Immunoblots utilizing the UL112 antibody identified seven distinct protein bands (p20, p26, p28, p34, p43, p50 and p84) expressed during the HCMV infectious cycle. After screening a cDNA library constructed from cells 72 h after infection with HCMV, only four different cDNA protein-producing constructs were obtained, and their ORFs corresponded to p34, p43, p50 and p84. The proteins p20, p26 and p28 were further shown to be selectively included within mature HCMV particles, virions, non-infectious enveloped particles and dense bodies. Immunoaffinity protein purification was used to prepare the samples for liquid chromatography coupled to tandem mass spectrometry. This analysis revealed that p20, p26 and p28 were derived from the UL112 ORF, most likely through post-translational proteolytic cleavage.

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“…Virions were isolated by glycerol-tartrate gradient centrifugation as described (39). 293T cells were cultured in Dulbecco's modified Eagle's medium (DMEM) (Gibco-BRL) supplemented with 10% FCS, glutamine, gentamicin, and 50 g of G418 (Geneticin; Gibco-BRL) per ml.…”

Section: Cells and Virusesmentioning

confidence: 99%

Identification of Glycoprotein gpTRL10 as a Structural Component of Human Cytomegalovirus

Spaderna

Blessing

Bogner

et al. 2002

J Virol

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Human cytomegalovirus (HCMV) has a coding capacity for glycoproteins which far exceeds that of other herpesviruses. Few of these proteins have been characterized. We have investigated the gene product(s) of reading frame 10, which is present in both the internal and terminal repeat regions of HCMV strain AD169 and only once in clinical isolates. The putative protein product is a 171-amino-acid glycoprotein with a theoretical mass of 20.5 kDa. We characterized the protein encoded by this reading frame in the laboratory strain AD169 and a recent isolate, TB40E. The results from both strains were comparable. Northern blot analyses showed that the gene was transcribed with early/late kinetics. Two proteins of 22 and 23.5-kDa were detected in virus-infected cells and in cells transiently expressing recombinant TRL10. Both forms contained only high-mannose-linked carbohydrate modifications. In addition, virus-infected cells expressed small amounts of the protein modified with complex N-linked sugars. Image analysis localized transiently expressed TRL10 to the endoplasmic reticulum. Immunoblot analyses as well as immunoelectron microscopy of purified virions demonstrated that TRL10 represents a structural component of the virus particle. Immunoblot analysis in the absence of reducing agents indicated that TRL10, like the other HCMV envelope glycoproteins, is present in a disulfide-linked complex. Sequence analysis of the TRL10 coding region in nine low-passage clinical isolates revealed strain-specific variation. In summary, the protein product of the TRL10 open reading frame represents a novel structural glycoprotein of HCMV and was termed gpTRL10.

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Human Cytomegalovirus: Purification of Enveloped Virions and Dense Bodies

Cited by 129 publications

References 0 publications

Characterization of enveloped tegument structures (L particles) produced by alphaherpesviruses: integrity of the tegument does not depend on the presence of capsid or envelope

Characterization of enveloped tegument structures (L particles) produced by alphaherpesviruses: integrity of the tegument does not depend on the presence of capsid or envelope

Novel virus-associated proteins encoded by UL112–113 of human cytomegalovirus

Identification of Glycoprotein gpTRL10 as a Structural Component of Human Cytomegalovirus

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