Intracellular Sequestration of the NKG2D Ligand ULBP3 by Human Cytomegalovirus

Bennett, Neil; Ashiru, Omodele; Morgan, Fiona J. E.; Pang, Yin Huei; Okecha, Georgina; Eagle, Robert A.; Trowsdale, John; Sissons, J. G. P.; Wills, Mark R.

doi:10.4049/jimmunol.1000789

Cited by 67 publications

(44 citation statements)

References 71 publications

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“…However, the evidence that in primary fibroblasts MICA was induced by both laboratory and low-passage HCMV strains suggests that the downmodulating activity exerted by the viral proteins UL142, US9, US18, and US20 on this ligand (14-17) was not sufficient to prevent its overall cell surface expression. Similarly, although UL142 was described to prevent expression of ULBP3 as well (67), in our settings this ligand was always increased, consistent with previous findings (57). These discrepancies may be related to different experimental conditions and/or to the considerable polymorphism in the UL142 sequence among different strains (68,69).…”

Section: Discussionsupporting

confidence: 79%

Distinct Roles for Human Cytomegalovirus Immediate Early Proteins IE1 and IE2 in the Transcriptional Regulation of MICA and PVR/CD155 Expression

Pignoloni

Fionda

Dell’Oste

et al. 2016

The Journal of Immunology

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Elimination of virus-infected cells by cytotoxic lymphocytes is triggered by activating receptors, among which NKG2D and DNAM-1/CD226 play an important role. Their ligands, that is, MHC class I–related chain (MIC) A/B and UL16-binding proteins (ULBP)1–6 (NKG2D ligand), Nectin-2/CD112, and poliovirus receptor (PVR)/CD155 (DNAM-1 ligand), are often induced on virus-infected cells, although some viruses, including human CMV (HCMV), can block their expression. In this study, we report that infection of different cell types with laboratory or low-passage HCMV strains upregulated MICA, ULBP3, and PVR, with NKG2D and DNAM-1 playing a role in NK cell–mediated lysis of infected cells. Inhibition of viral DNA replication with phosphonoformic acid did not prevent ligand upregulation, thus indicating that early phases of HCMV infection are involved in ligand increase. Indeed, the major immediate early (IE) proteins IE1 and IE2 stimulated the expression of MICA and PVR, but not ULBP3. IE2 directly activated MICA promoter via its binding to an IE2-responsive element that we identified within the promoter and that is conserved among different alleles of MICA. Both IE proteins were instead required for PVR upregulation via a mechanism independent of IE DNA binding activity. Finally, inhibiting IE protein expression during HCMV infection confirmed their involvement in ligand increase. We also investigated the contribution of the DNA damage response, a pathway activated by HCMV and implicated in ligand regulation. However, silencing of ataxia telangiectasia mutated, ataxia telangiectasia and Rad3–related protein, and DNA-dependent protein kinase did not influence ligand expression. Overall, these data reveal that MICA and PVR are directly regulated by HCMV IE proteins, and this may be crucial for the onset of an early host antiviral response.

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Section: Discussionsupporting

confidence: 79%

Distinct Roles for Human Cytomegalovirus Immediate Early Proteins IE1 and IE2 in the Transcriptional Regulation of MICA and PVR/CD155 Expression

Pignoloni

Fionda

Dell’Oste

et al. 2016

The Journal of Immunology

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“…Interestingly, all OWM CMVs lack several known HCMV antagonists of NK cell function, including UL16, which retains the NKG2D ligands MICB, ULBP1, and ULBP2 (22,77), UL141, which retains the NKG2D ligands MICA and ULBP3 (4,8,11), and UL18, which encodes an MHC-I homologue that binds to the inhibitory NK cell receptor LIR-1 (16,56,62). Since evasion of NK cell immunity is conserved in MCMV (6), it is likely that OWM CMV-specific genes will encode NK cell evasins.…”

Section: Resultsmentioning

confidence: 99%

Reevaluation of the Coding Potential and Proteomic Analysis of the BAC-Derived Rhesus Cytomegalovirus Strain 68-1

Malouli

Nakayasu

Viswanathan

et al. 2012

J Virol

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Cytomegaloviruses are highly host restricted, resulting in cospeciation with their hosts. As a natural pathogen of rhesus macaques (RM), rhesus cytomegalovirus (RhCMV) has therefore emerged as a highly relevant experimental model for pathogenesis and vaccine development due to its close evolutionary relationship to human CMV (HCMV). Most in vivo experiments performed with RhCMV employed strain 68-1 cloned as a bacterial artificial chromosome (BAC). However, the complete genome sequence of the 68-1 BAC has not been determined. Furthermore, the gene content of the RhCMV genome is unknown, and previous open reading frame (ORF) predictions relied solely on uninterrupted ORFs with an arbitrary cutoff of 300 bp. To obtain a more precise picture of the actual proteins encoded by the most commonly used molecular clone of RhCMV, we reevaluated the RhCMV 68-1 BAC genome by whole-genome shotgun sequencing and determined the protein content of the resulting RhCMV virions by proteomics. By comparing the RhCMV genome to those of several related Old World monkey (OWM) CMVs, we were able to filter out many unlikely ORFs and obtain a simplified map of the RhCMV genome. This comparative genomics analysis suggests a high degree of ORF conservation among OWM CMVs, thus decreasing the likelihood that ORFs found only in RhCMV comprise true genes. Moreover, virion proteomics independently validated the revised ORF predictions, since only proteins that were conserved across OWM CMVs could be detected. Taken together, these data suggest a much higher conservation of genome and virion structure between CMVs of humans, apes, and OWMs than previously assumed.

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“…Compared with the genome of the prototype laboratory strain, AD169, genomes of low-passage HCMV clinical isolates contain the UL/b' region, including ORFs UL133-UL151, considered as a critical candidate cluster to clinical pathogenesis (3). Although the UL/b' region is not essential to viral growth or replication (4), products of this region, including UL141, UL142 and UL144 have been experimentally identified to aid in viral escape from immune surveillance through interactions with cellular molecules (5)(6)(7)(8)(9)(10).…”

Section: Introductionmentioning

confidence: 99%

Identification of immediate early gene X-1 as a cellular target gene of hcmv-mir-UL148D

Wang

Huang

et al. 2013

International Journal of Molecular Medicine

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Abstract. Human cytomegalovirus (HCMV) is a herpesvirus that causes congenital diseases and opportunistic infections in immunocompromised individuals. Its functional proteins and microRNAs (miRNAs) facilitate efficient viral propagation by altering host cell behavior. The identification of functional target genes of miRNAs is an important step in the study of HCMV pathogenesis. HCMV encodes at least 14 miRNAs, including hcmv-mir-UL148D, which resides in the HCMV UL/b' region. hcmv-mir-UL148D is the only miRNA encoded by the HCMV UL/b' region; however, its targets and functional effects have not yet been eludidated. In this study, hybrid-PCR screening was used to identify target genes and dual luciferase reporter assay was used to evaluate the binding effect of hcmv-miR-UL148D to the 3' untranslated region (3'UTR) of IEX-1. In addition, western blot analysis was used to detect the expression kinetics of IEX-1 protein and apoptosis assay was used to identify the effects of hcmv-miR-UL148D on cell apoptosis. The hybrid-PCR results showed that only one binding site in the 3'UTR of the cellular gene, human immediate early gene X-1 (IEX-1), was completely complementary to an 11 nucleotide (nt) sequence in the 5' terminus of hcmvmir-UL148D, including the entire seed region. The binding site was demonstrated to be functional by dual luciferase reporter assay with a 47% repression of the relative luciferase activity. In an in vitro system of human embryonic kidney 293 (HEK293) cells, the ectopically expressed hcmv-mir-UL148D exhibited a downregulatory effect on IEX-1 expression, and decreased the cell apoptosis induced by transfected IEX-1. Our data demonstrate that hcmv-mir-UL148D targets the cellular gene, IEX-1, downregulating its expression and thus results in anti-apoptotic effects.

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Intracellular Sequestration of the NKG2D Ligand ULBP3 by Human Cytomegalovirus

Cited by 67 publications

References 71 publications

Distinct Roles for Human Cytomegalovirus Immediate Early Proteins IE1 and IE2 in the Transcriptional Regulation of MICA and PVR/CD155 Expression

Distinct Roles for Human Cytomegalovirus Immediate Early Proteins IE1 and IE2 in the Transcriptional Regulation of MICA and PVR/CD155 Expression

Reevaluation of the Coding Potential and Proteomic Analysis of the BAC-Derived Rhesus Cytomegalovirus Strain 68-1

Identification of immediate early gene X-1 as a cellular target gene of hcmv-mir-UL148D

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