Genetic content of wild-type human cytomegalovirus

Dolan, Aidan; Cunningham, Charles; Hector, Ralph D.; Hassan‐Walker, Aycan F.; Lee, Lydia; Addison, Clare; Dargan, Derrick J.; McGeoch, Duncan J.; Gatherer, Derek; Emery, Vincent C.; Griffiths, Paul; Sinzger, Christian; McSharry, Brian P.; Wilkinson, Gavin William Grahame; Davison, Andrew J.

doi:10.1099/vir.0.79888-0

Cited by 508 publications

(516 citation statements)

References 45 publications

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“…Interestingly, a similar unanticipated deletion was found in the HCMV FIXBAC (Murphy et al, 2003b), which was constructed using the same BAC vector and flanking homologous arms (Hahn et al, 2002). With regard to previously reported TB40/E-variants, TB40-BAC4 resembles the Bart strain in that UL141 has a frameshift insertion at codon 63 (Tomasec et al, 2005); however, unlike strain Bart, UL144 and UL145 are intact and thus TB40-BAC4 is identical to the TB40/E sequence published by Dolan et al (2004) in all three genes. The entire TB40-BAC4 sequence was annotated in analogy to other previously annotated HCMV strains and has been deposited under the designation TB40-BAC4 at the GenBank database (accession no.…”

Section: Sequence Alignment Of Tb40-bac4 With Various Hcmv Genomesmentioning

confidence: 96%

Cloning and sequencing of a highly productive, endotheliotropic virus strain derived from human cytomegalovirus TB40/E

Sinzger

Hahn

Digel

et al. 2008

Journal of General Virology

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343

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Human cytomegalovirus (HCMV) strain TB40/E, replicates efficiently, exhibits a broad cell tropism and is widely used for infection of endothelial cells and monocyte-derived cells yet has not been available in a phenotypically homogeneous form compatible with genetic analysis. To overcome this problem, we cloned the TB40/E strain into a bacterial artificial chromosome (BAC) vector. Both highly endotheliotropic and poorly endotheliotropic virus clones, representing three distinct restriction fragment patterns, were reconstituted after transfection of BAC clones derived from previously plaque-purified strain TB40/E. For one of the highly endotheliotropic clones, TB40-BAC4, we provide the genome sequence. Two BACs with identical restriction fragment patterns but different cell tropism were further analysed in the UL128-UL131A gene region. Sequence analysis revealed one coding-relevant adenine insertion at position 332 of UL128 in the BAC of the poorly endotheliotropic virus, which caused a frameshift in the C-terminal part of the coding sequence. Removal of this insertion by markerless mutagenesis restored the highly endotheliotropic phenotype, indicating that the loss of endothelial cell tropism was caused by this insertion. In conclusion, HCMV strain TB40/E, which combines the high endothelial cell tropism of a clinical isolate with the high titre growth of a cell culture adapted strain, is now available as a BAC clone suitable for genetic engineering. The results also suggest BAC cloning as a suitable method for selection of genetically defined virus clones.

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Section: Sequence Alignment Of Tb40-bac4 With Various Hcmv Genomesmentioning

confidence: 96%

Cloning and sequencing of a highly productive, endotheliotropic virus strain derived from human cytomegalovirus TB40/E

Sinzger

Hahn

Digel

et al. 2008

Journal of General Virology

Self Cite

343

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“…The reasons for these conflicting results remain unresolved. In contrast to endotheliotropic/clinical HCMV strains, such as Towne/E and TB40/E, laboratory/ fibroblast-adapted strains, such as AD169, Towne, and TB40/F, have reduced infectivity of endothelial/epithelial cells and monocytes because of losses or mutations in the U L BЈ region of the viral genome (35)(36)(37). Expression of distinct viral glycoproteins from the U L BЈ region allows viral infection of endothelial/epithelial cells but not of fibroblasts, indicating the existence of cell type-specific receptors for different strains of HCMV (38).…”

Section: Discussionmentioning

confidence: 99%

Activation of EGFR on monocytes is required for human cytomegalovirus entry and mediates cellular motility

Chan

Nogalski²,

Yurochko³

2009

Proc. Natl. Acad. Sci. U.S.A.

169

287

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Human cytomegalovirus (HCMV) rapidly induces a mobile and functionally unique proinflammatory monocyte following infection that is proposed to mediate viral spread. The cellular pathways used by HCMV to initiate these biological changes remain unknown. Here, we document the expression of the epidermal growth factor receptor (EGFR) on the surface of human peripheral blood monocytes but not on other blood leukocyte populations. Inhibition of EGFR signaling abrogated viral entry into monocytes, indicating that EGFR can serve as a cellular tropism receptor. Moreover, HCMV-activated EGFR was required for the induction of monocyte motility and transendothelial migration, two biological events required for monocyte extravasation into peripheral tissue, and thus viral spread. Transcriptome analysis revealed that HCMV-mediated EGFR signaling up-regulated neural Wiskott-Aldrich syndrome protein (N-WASP), an actin nucleator whose expression and function are normally limited in leukocytes. Knockdown of N-WASP expression blocked HCMV-induced but not phorbol 12-myristate 13-acetate (PMA)-induced monocyte motility, suggesting that a switch to and/or the distinct use of a new actin nucleator controlling motility occurs during HCMV infection of monocytes. Together, these data provide evidence that EGFR plays an essential role in the immunopathobiology of HCMV by mediating viral entry into monocytes and stimulating the aberrant biological activity that promotes hematogenous dissemination.T he wide range of pathological complications associated with human cytomegalovirus (HCMV) infection is a direct consequence of viral spread to peripheral organ sites and the broad cellular tropism of the virus (1). Monocytes are primary in vivo targets for HCMV and are believed to be responsible for hematogenous dissemination of HCMV to multiple organ systems (2). We previously showed that HCMV infection of monocytes polarized the infected cell toward a distinct proinflammatory phenotype that possessed the distinct biological changes necessary to promote viral spread (3-5). Specifically, HCMV infection induced polarization of infected monocytes toward an M1 proinflammatory cell type that simultaneously exhibited characteristics associated with an M2 antiinflammatory macrophage (6). The infected monocytes also displayed a high level of chemokinesis when compared with monocytes activated by LPS or phorbol 12-myristate 13-acetate (PMA) (4, 5). Moreover, an accelerated rate of differentiation from short-lived monocytes (nonpermissive for HCMV replication) into long-lived macrophages (permissive for HCMV replication) was observed following infection with HCMV (4). Based on our studies, we suggest that during primary infection, newly infected peripheral monocytes acquire a motile phenotype that promotes exit of the infected cell from the circulating blood into multiple organ tissue despite the absence of a chemotactic gradient. Once in the surrounding tissue, differentiation into HCMV replication-competent macrophages occurs, resulting in viral spread...

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“…The genomic complexity of HCMV is still being addressed. Primary strains of HCMV contain several more ORFs than laboratory strains of HCMV (Dolan et al, 2004). Also, recent highresolution transcriptome mapping (Gatherer et al, 2011) and ribosome profiling (Stern-Ginossar et al, 2012) have revealed the presence of many previously unrecognized viral RNA transcripts and non-canonical ORFs.…”

Section: Further Questions and Future Perspectivesmentioning

confidence: 99%

“…pp71 degrades hDaxx by a proteasome-dependent, ubiquitin-independent mechanism (Kalejta & Shenk, 2003) and displaces ATRX from PML-NBs (Lukashchuk et al, 2008). The presence of IE1 promotes the disruption of PML-NBs, induces the proteasome-independent loss of SUMOylated PML and Sp100 proteins (Ahn & Hayward, 1997;Kang et al, 2006;Kim et al, 2011;Korioth et al, 1996;Lee et al, 2004;Wilkinson et al, 1998;Xu et al, 2001) and potentially induces the proteasome-dependent degradation of unSUMOylated Sp100 late in infection . Importantly, IE1 does not act like its well-studied counterpart expressed by HSV, ICP0.…”

Section: Introductionmentioning

confidence: 99%

Viral and cellular subnuclear structures in human cytomegalovirus-infected cells

Strang

2015

Journal of General Virology

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In human cytomegalovirus (HCMV)-infected cells, a dramatic remodelling of the nuclear architecture is linked to the creation, utilization and manipulation of subnuclear structures. This review outlines the involvement of several viral and cellular subnuclear structures in areas of HCMV replication and virus-host interaction that include viral transcription, viral DNA synthesis and the production of DNA-filled viral capsids. The structures discussed include those that promote or impede HCMV replication (such as viral replication compartments and promyelocytic leukaemia nuclear bodies, respectively) and those whose role in the infected cell is unclear (for example, nucleoli and nuclear speckles). Viral and cellular proteins associated with subnuclear structures are also discussed. The data reviewed here highlight advances in our understanding of HCMV biology and emphasize the complexity of HCMV replication and virus-host interactions in the nucleus. IntroductionThe betaherpesvirus human cytomegalovirus (HCMV) is a widespread opportunistic pathogen that severely affects immunocompromised and immunodeficient populations (Mocarski et al., 2007). Like all herpesviruses, HCMV undergoes either productive or latent infection. During productive infection, infectious virus is produced, while in latent infection the virus becomes largely transcriptionally quiescent (Mocarski et al., 2007). Like other herpesviruses, many events that are critical for productive HCMV replication take place within the nucleus. These include essential steps in viral replication, such as: the transcriptional cascade of immediate-early (IE), early and late viral RNA transcripts, synthesis of viral DNA and the production of DNA-containing capsids (Mocarski et al., 2007). Compared with the prototype human herpesvirus, the alphaherpesvirus herpes simplex virus (HSV), certain features of productive HCMV infection are not well characterized. However, it is clear that several subnuclear structures are involved in HCMV genome replication and virus-host interactions. Uncovering the roles of these structures has led to significant advances in our understanding of HCMV biology. This review summarizes the involvement of several viral and cellular subnuclear structures in productive HCMV infection. The participation of each structure in HCMV replication and virus-host interactions is described from entry of the viral genome into the nucleus to the egress of viral capsids through the nuclear membrane. The data discussed highlight recent advances in our understanding of subnuclear structure function, introduce viral and cellular factors associated with subnuclear structures and indicate what questions have yet to be answered.Entry of the HCMV genome into the nucleus: the nuclear pore complex, nuclear speckles, promyelocytic leukaemia protein nuclear bodies and pp150 bodies HCMV genome entry into the nucleus is poorly defined but may be similar to movement of the HSV DNA genome through the nuclear pore complex (NPC) (Kobiler et al., 2012) (Fig. 1a). HCMV capsid ...

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Genetic content of wild-type human cytomegalovirus

Cited by 508 publications

References 45 publications

Cloning and sequencing of a highly productive, endotheliotropic virus strain derived from human cytomegalovirus TB40/E

Cloning and sequencing of a highly productive, endotheliotropic virus strain derived from human cytomegalovirus TB40/E

Activation of EGFR on monocytes is required for human cytomegalovirus entry and mediates cellular motility

Viral and cellular subnuclear structures in human cytomegalovirus-infected cells

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