Analysis of the nucleotide sequence of the guinea pig cytomegalovirus (GPCMV) genome

bPML nuclear bodies and their associated functions are part of an intrinsic cellular mechanism aimed at maintaining transcriptional control over viral gene expression and preventing replication of invading viruses. To overcome these barriers, many viruses express early nonstructural, multifunctional proteins to support the viral replication cycle or modulate host immune responses. Virion proteins constituting the invading particle are traditionally investigated for their role in transport during entry or egress and in the assembly of new virions. The additional functions of virion proteins have largely been ignored, in contrast to those of their nonstructural counterparts. A number of recent reports suggest that several virion proteins may also play vital roles in gene activation processes, in particular by counteracting intrinsic immune mechanisms mediated by the PML nuclear body-associated cellular factors Daxx, ATRX, and Sp100. These virion proteins share several features with their more potent nonstructural counterparts, and they may serve to bridge the gap in the early phase of an infection until immediate early viral gene expression is established. In this review, we discuss how virion proteins are an integral part of gene regulation among several viral families and to what extent structural proteins of incoming virions may contribute to species barrier, latency, and oncogenesis.W hen viral genetic information is transferred from cell to cell, it must be compacted and transcriptionally inactivated for storage in the viral capsid during transport. This process needs to be actively reversed upon infection against cellular transcriptional repression through intrinsic antiviral mechanisms. An important factor in the cellular control of viral gene expression is death domain-associated protein (Daxx). Daxx was initially described as a modulator of apoptotic signaling (1), but recent reports suggest that Daxx plays an active role in gene regulation by repressing or modulating transcription through chromatin remodeling (2, 3). Daxx mainly cooperates with alpha-thalassemia retardation syndrome x-linked (ATRX), a putative member of the SNF2 family of ATP-dependent chromatin-remodeling proteins. In this repressive complex, ATRX acts as the core ATPase subunit, while Daxx is the targeting factor, leading to histone deacetylase (HDAC) recruitment (4-6).Daxx is found associated with the promyelocytic leukemia protein nuclear body (PML-NB) or chromatin. Association with PML-NB alleviates gene repression and activates apoptosis, while chromatin-bound Daxx represses transcription (7-9). It has been long established that PML-NBs are nuclear structures with antiviral activity, accumulating an expanding number of transient or constitutive cellular factors involved in transcriptional control, chromatin remodeling, genome integrity, apoptosis, and tumor suppression (10). Moreover, several PML-NB constituents can be induced by type I and II interferon (IFN), resulting in increased antiviral activity by attenuating viral gene exp...

Section: Daxx Impairment By Virion Proteins: Overcoming the Species Bsupporting

confidence: 62%

Virion Factors That Target Daxx To Overcome Intrinsic Immunity

Schreiner

Wodrich

2013

“…The new ORFs Rh00.2, Rh94.1, Rh96.1, Rh228.1, and Rh231 are predicted for both RhCMV strains, whereas ORFs Rh163.1, Rh165.1, and Rh166.1 are conserved between RhCMV 68-1 and the ULb= region of the low-passage-number isolate RhCMV CNPRC as described previously (51). However, the total number of ORFs predicted by this method for either RhCMV strain was substantially higher than that for any other published CMV genome (HCMV [25], CCMV [19], mouse CMV [MCMV] [60], rat CMV [RCMV] [76], and guinea pig CMV [GPCMV] [37,67]). Moreover, this annotation predicted that eight ORFs were unique to RhCMV 68-1 (Rh09, Rh39, Rh61, Rh93, Rh94, Rh142.4, Rh153, and Rh220.1) and that four ORFs were unique to RhCMV 180.92 (Rh13.1, Rh106.1, Rh142.3, and Rh178.2).…”

Section: Resultsmentioning

confidence: 87%

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

Malouli

Nakayasu

Viswanathan

et al. 2012

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.

“…A major drawback in GPCMV research has largely been overcome by the recent sequencing of the viral genome and the development of infectious bacterial artificial chromosome (BAC) clones of GPCMV (15,(26)(27)(28)(29). Manipulation of an infectious GPCMV BAC has allowed the preliminary study of some viral genes (1,(30)(31)(32)(33)(34)(35)(36).…”

mentioning

confidence: 99%

“…Manipulation of an infectious GPCMV BAC has allowed the preliminary study of some viral genes (1,(30)(31)(32)(33)(34)(35)(36). Analysis of the viral genome (15,29) indicated that GPCMV encodes homologs to the HCMV glycoproteins (gB, gH, gL, gM, gN, and gO) in genes colinear with the HCMV genome (designated GP55, GP75, GP115, GP100, GP73, and GP74, respectively). In HCMV, these six glycoproteins (gB, gH, gL, gM, gN, and gO) are required for fibroblast cell entry, and they form the glycoprotein complexes gCI (gB), gCII (gM/gN), and gcIII (gH/gL/gO) on the viral membrane (37)(38)(39).…”

mentioning

confidence: 99%

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A Novel Non-Replication-Competent Cytomegalovirus Capsid Mutant Vaccine Strategy Is Effective in Reducing Congenital Infection

Choi

Root

McGregor

2016

Self Cite

Congenital cytomegalovirus (CMV) infection is a leading cause of mental retardation and deafness in newborns. The guinea pig is the only small animal model for congenital CMV infection. A novel CMV vaccine was investigated as an intervention strategy against congenital guinea pig cytomegalovirus (GPCMV) infection. In this disabled infectious single-cycle (DISC) vaccine strategy, a GPCMV mutant virus was used that lacked the ability to express an essential capsid gene (the UL85 homolog GP85) except when grown on a complementing cell line. In vaccinated animals, the GP85 mutant virus (GP85 DISC) induced an antibody response to important glycoprotein complexes considered neutralizing target antigens (gB, gH/gL/gO, and gM/gN). The vaccine also generated a T cell response to the pp65 homolog (GP83), determined via a newly established guinea pig gamma interferon enzyme-linked immunosorbent spot assay. In a congenital infection protection study, GP85 DISC-vaccinated animals and a nonvaccinated control group were challenged during pregnancy with wild-type GPCMV (10 5 PFU). The pregnant animals carried the pups to term, and viral loads in target organs of pups were analyzed. Based on live pup births in the vaccinated and control groups (94.1% versus 63.6%), the vaccine was successful in reducing mortality (P ‫؍‬ 0.0002). Additionally, pups from the vaccinated group had reduced CMV transmission, with 23.5% infected target organs versus 75.9% in the control group. Overall, these preliminary studies indicate that a DISC CMV vaccine strategy has the ability to induce an immune response similar to that of natural virus infection but has the increased safety of a non-replication-competent virus, which makes this approach attractive as a CMV vaccine strategy. IMPORTANCECongenital CMV infection is a leading cause of mental retardation and deafness in newborns. An effective vaccine against CMV remains an elusive goal despite over 50 years of CMV research. The guinea pig, with a placenta structure similar to that in humans, is the only small animal model for congenital CMV infection and recapitulates disease symptoms (e.g., deafness) in newborn pups. In this report, a novel vaccine strategy against congenital guinea pig cytomegalovirus (GPCMV) infection was developed, characterized, and tested for efficacy. This disabled infectious single-cycle (DISC) vaccine strategy induced a neutralizing antibody or a T cell response to important target antigens. In a congenital infection protection study, animals were protected against CMV in comparison to the nonvaccinated group (52% reduction of transmission). This novel vaccine was more effective than previously tested gB-based vaccines and most other strategies involving live virus vaccines. Overall, the DISC vaccine is a safe and promising approach against congenital CMV infection. H uman cytomegalovirus (HCMV), a betaherpesvirus, has evolved very closely with its human host. Virus infection in a healthy host is normally asymptomatic but leads to a lifelong infection. In contrast,...