Proteomic Analyses of Human Cytomegalovirus Strain AD169 Derivatives Reveal Highly Conserved Patterns of Viral and Cellular Proteins in Infected Fibroblasts

The degree of relatedness between previously identified cross-hybridizing regions within human cytomegalovirus strain AD169 and the avian retrovirus oncogene v-myc were investigated by nucleotide sequence comparison. We found that the homologous regions between the human cytomegalovirus genome and v-myc are limited to short G + C-rich regions in each genome and that the human cytomegalovirus genome shares little or no homology with the human c-myc gene.

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Human Cytomegalovirus TR Strain Glycoprotein O Acts as a Chaperone Promoting gH/gL Incorporation into Virions but Is Not Present in Virions

Journal of Virology ◽

10.1128/jvi.02256-09 ◽

2009 ◽

Vol 84 (5) ◽

pp. 2597-2609 ◽

Cited By ~ 38

Author(s):

Brent J. Ryckman ◽

Marie C. Chase ◽

David C. Johnson

Keyword(s):

Endothelial Cells ◽

Human Cytomegalovirus ◽

Null Mutant ◽

Virus Particles ◽

Extracellular Virus ◽

Biochemical Studies ◽

Infected Cells ◽

Low Passage ◽

Er Export ◽

Strain Ad169

ABSTRACT Human cytomegalovirus (HCMV) produces the following two gH/gL complexes: gH/gL/gO and gH/gL/UL128-131. Entry into epithelial and endothelial cells requires gH/gL/UL128-131, and we have provided evidence that gH/gL/UL128-131 binds saturable epithelial cell receptors to mediate entry. HCMV does not require gH/gL/UL128-131 to enter fibroblasts, and laboratory adaptation to fibroblasts results in mutations in the UL128-131 genes, abolishing infection of epithelial and endothelial cells. HCMV gO-null mutants produce very small plaques on fibroblasts yet can spread on endothelial cells. Thus, one prevailing model suggests that gH/gL/gO mediates infection of fibroblasts, while gH/gL/UL128-131 mediates entry into epithelial/endothelial cells. Most biochemical studies of gO have involved the HCMV lab strain AD169, which does not assemble gH/gL/UL128-131 complexes. We examined gO produced by the low-passage clinical HCMV strain TR. Surprisingly, TR gO was not detected in purified extracellular virus particles. In TR-infected cells, gO remained sensitive to endoglycosidase H, suggesting that the protein was not exported from the endoplasmic reticulum (ER). However, TR gO interacted with gH/gL in the ER and promoted export of gH/gL from the ER to the Golgi apparatus. Pulse-chase experiments showed that a fraction of gO remained bound to gH/gL for relatively long periods, but gO eventually dissociated or was degraded and was not found in extracellular virions or secreted from cells. The accompanying report by P. T. Wille et al. (J. Virol., 84:2585-2596, 2010) showed that a TR gO-null mutant failed to incorporate gH/gL into virions and that the mutant was unable to enter fibroblasts and epithelial and endothelial cells. We concluded that gO acts as a molecular chaperone, increasing gH/gL ER export and incorporation into virions. It appears that gO competes with UL128-131 for binding onto gH/gL but is released from gH/gL, so that gH/gL (lacking UL128-131) is incorporated into virions. Thus, our revised model suggests that both gH/gL and gH/gL/UL128-131 are required for entry into epithelial and endothelial cells.

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Identification of Glycoprotein gpTRL10 as a Structural Component of Human Cytomegalovirus

Journal of Virology ◽

10.1128/jvi.76.3.1450-1460.2002 ◽

2002 ◽

Vol 76 (3) ◽

pp. 1450-1460 ◽

Cited By ~ 13

Author(s):

S. Spaderna ◽

H. Blessing ◽

E. Bogner ◽

W. Britt ◽

M. Mach

Keyword(s):

Human Cytomegalovirus ◽

Structural Component ◽

Laboratory Strain ◽

Immunoblot Analysis ◽

Protein Product ◽

Clinical Isolates ◽

Coding Region ◽

Reading Frame ◽

Infected Cells ◽

Strain Ad169

ABSTRACT 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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Impact of new cyclooxygenase 2 inhibitors on human cytomegalovirus replication in vitro

Antiviral Therapy ◽

10.1177/13596535211064078 ◽

2021 ◽

pp. 135965352110640

Author(s):

D Andouard ◽

R Gueye ◽

S Hantz ◽

C Fagnère ◽

B Liagre ◽

...

Keyword(s):

Human Cytomegalovirus ◽

Hcmv Infection ◽

Laboratory Strain ◽

Cyclooxygenase 2 ◽

Immunocompromised Patients ◽

Toxic Compound ◽

Cox 2 ◽

Cyclooxygenase 2 Inhibitors ◽

Strain Ad169

Background Human cytomegalovirus (HCMV) is involved in complications on immunocompromised patients. Current therapeutics are associated with several drawbacks, such as nephrotoxicity. Purpose: As HCMV infection affects inflammation pathways, especially prostaglandin E2 (PGE2) production via cyclooxygenase 2 enzyme (COX-2), we designed 2'-hydroxychalcone compounds to inhibit human cytomegalovirus. Study design We first selected the most efficient new synthetic chalcones for their effect against COX-2-catalyzed PGE2. Study sample Among the selected compounds, we assessed the antiviral efficacy against different HCMV strains, such as the laboratory strain AD169 and clinical strains (naïve or multi-resistant to conventional drugs) and toxicity on human cells. Results The most efficient and less toxic compound (chalcone 7) was tested against HCMV in combination with other antiviral molecules: artesunate (ART), baicalein (BAI), maribavir (MBV), ganciclovir (GCV), and quercetin (QUER) using Compusyn software. Association of chalcone 7 with MBV and BAI is synergistic, antagonistic with QUER, and additive with GCV and ART. Conclusion These results provide a promising search path for potential bitherapies against HCMV.

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Identification of Proteins in Human Cytomegalovirus (HCMV) Particles: the HCMV Proteome

Journal of Virology ◽

10.1128/jvi.78.20.10960-10966.2004 ◽

2004 ◽

Vol 78 (20) ◽

pp. 10960-10966 ◽

Cited By ~ 408

Author(s):

Susan M. Varnum ◽

Daniel N. Streblow ◽

Matthew E. Monroe ◽

Patricia Smith ◽

Kenneth J. Auberry ◽

...

Keyword(s):

Human Cytomegalovirus ◽

Viral Proteins ◽

Capillary Liquid Chromatography ◽

Open Reading Frames ◽

Cellular Proteins ◽

Ion Cyclotron Resonance ◽

Virion Protein ◽

Enveloped Virus ◽

Dense Bodies ◽

Large Complex

ABSTRACT Human cytomegalovirus (HCMV), a member of the herpesvirus family, is a large complex enveloped virus composed of both viral and cellular gene products. While the sequence of the HCMV genome has been known for over a decade, the full set of viral and cellular proteins that compose the HCMV virion are unknown. To approach this problem we have utilized gel-free two-dimensional capillary liquid chromatography-tandem mass spectrometry (MS/MS) and Fourier transform ion cyclotron resonance MS to identify and determine the relative abundances of viral and cellular proteins in purified HCMV AD169 virions and dense bodies. Analysis of the proteins from purified HCMV virion preparations has indicated that the particle contains significantly more viral proteins than previously known. In this study, we identified 71 HCMV-encoded proteins that included 12 proteins encoded by known viral open reading frames (ORFs) previously not associated with virions and 12 proteins from novel viral ORFs. Analysis of the relative abundance of HCMV proteins indicated that the predominant virion protein was the pp65 tegument protein and that gM rather than gB was the most abundant glycoprotein. We have also identified over 70 host cellular proteins in HCMV virions, which include cellular structural proteins, enzymes, and chaperones. In addition, analysis of HCMV dense bodies indicated that these viral particles are composed of 29 viral proteins with a reduced quantity of cellular proteins in comparison to HCMV virions. This study provides the first comprehensive quantitative analysis of the viral and cellular proteins that compose infectious particles of a large complex virus.

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