Convergent capture of retroviral superantigens by mammalian herpesviruses : Nature Communications : Nature Publishing Group

Convergent capture of retroviral superantigens by mammalian herpesviruses : Nature Communications : Nature Publishing Group

The present invention relates to the use of certain derivatives of acyclovir, penciclovir and ganciclovir, and other anti-herpetic compounds to prevent prodrome and vesicle outbreaks in a human subject having a herpes virus infection, particularly HSV-II. KSHV latency genes (LANA-1, v-FLIP, v-cyclin, K8 and K12) were transduced (~90% efficiency; Supplementary Figure S1Aa and b) in HMVEC-d cells using a lentiviral transduction method. latent infection in which most of the viral genome is transcriptionally silent. BDCRB blocks the processing and maturation of viral DNA, whereas 1263W94 inhibits the viral enzyme pUL97 and interferes with DNA synthesis. In TLE, the filtering function of the dentate gyrus is impaired as a result of the selective loss of inhibitory interneurons and because of synaptic reorganization, including the sprouting of dentate granule cell axons (i.e., mossy fibers) that causes the formation of positive-feedback reverberating excitatory circuits. In addition, we found VP22 is required for the accumulation of a subset of mRNAs to wild-type levels at early, but not late, times in infection. Rice, J.

Biological systems can share a common mechanism either because of descent from a common ancestral system or molecule (termed homology), or because of convergent evolution of two unrelated systems or molecules (termed analogy). These findings suggest that ICP0 mRNA synthesis is differentially regulated in HSV-infected cells by the virus-encoded repressor activity embedded in ICP4, and a virus-encoded antirepressor, ICP0. Viral adaptations to new hosts primarily manifest as amino acid substitutions which can allow more efficient virus cell entry into the new host [3,4], block interactions with detrimental host proteins [5,6] or promote escape from both the new and the old host’s immune responses [7,8]. So far, limited attention has been paid to the cellular determinants that route HSV to one or another entry pathway. Heterologous immunity appears to be a common phenomenon among closely related pathogens, for example, different strains of influenza or DENV, or among different members of the same virus group, such as hantaviruses, arenaviruses, and flaviviruses [2]. The history of the visualization of intrinsic resistance factors during herpesvirus infection dates back to early studies by Gerd Maul and colleagues, who discovered that a group of then uncharacterized cellular proteins form distinct nuclear foci. (a) Cartoon of a partial genome alignment of SaHV2, AtHV3 and RHVP, showing conserved gene blocks in blank boxes and lineage-specific genes in colour, labeled according to product name.

The figure also shows an alignment for SaHV2 and AtHV3 of the region between ORF11 and the minor capsid gene. The red graph depicts the percentage nucleotide-level similarity shared by the viruses. (b) Maximum-likelihood phylogenies of host IL-17 and CD59 from primates including a viral homologue in SaHV2. SaHV2 independently acquired these genes after diverging from AtHV3. The gene for CD59 in particular, appears to have been captured from the squirrel monkeys, which are the current host of the virus. Only the primate tips have been made visible. The scale bar represents the number of nucleotide substitutions per site.

The results of 1,000 nonparametric bootstrap replicates are indicated at each node. Primary infection (airborne) due to herpes varicella-zoster usually affects preschool children, causing chickenpox, with rare complications usually affecting the immunocompromised host. Small RNAs derived from this sample were used to prepare cDNAs for 454 sequencing13. 1 and Supplementary Fig. Epstein-Barr Virus (EBV, also denoted human herpes virus 4) infection results in activation of the RAG genes required for V(D)J recombination [17]–[20], suggesting that the viral and host genes are part of a similar regulatory network. Gammaherpesviruses include the human pathogens Epstein-Barr virus (EBV/HHV-4) and Kaposi’s sarcoma-associated herpesvirus (KSHV/HHV-8) [8], the etiologic agents of some of the most common AIDS-associated cancers, such as Burkitt’s lymphoma, nasopharyngeal carcinoma, and primary effusion lymphoma. Also, with Elephant Endotheliotropic Herpesvirus (EEHV) a common health issue for Elephants both in the care of zoos and in the wild, the Saint Louis Zoo has been instrumental in pursuing the latest EEHV detection and testing protocols.

2). The natural host range of individual viruses is usually restricted to a single species. We also searched for betaretrovirus ERVs using MMTV polymerase (Pol) as the query, to identify those closely related to MMTV that did not show sag similarity. To investigate the evolution of herpesvirus sag acquisition, we used the collected sequences to conduct a series of phylogenetic analyses that included data from both sets of searches. No. A conserved 100 amino-acid region of the sag sequences of SaHV2, AtHV3, RHVP and MMTV was used as a query. ).

Furthermore, 1263W94 inhibits the phosphorylation and accumulation of EBV early antigen D, an essential cofactor in EBV replication (11). For the basic scientist, Borna disease virus provides yet another valuable tool to study the functional organization of the hippocampus and the role of various components that regulate the excitability of its circuitry as well as providing a means to learn more about the mechanisms of TLE. Vero and V49 cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 4.0 mM l-glutamine, 4.5 g/liter glucose, 125 units/ml penicillin, 0.125 mg/ml streptomycin, and either 10% newborn calf serum or 10% fetal bovine serum, respectively. In general, the ICP27 homologs that have been studied are similar to ICP27 in that they stimulate viral gene expression, and in some cases this can be shown to occur at a posttranscriptional level (13, 22, 42, 45, 77, 78). The genome and termini of EBV have enriched G/C content (approximately 70% G/C), thus regions resembling a V(D)J RSS nanomer (A/T rich regions that contribute to DNA bending and interactions with bending proteins) are relatively uncommon. was required to observe repression of ICP0 mRNA synthesis in HSV-infected cells; 2. The family also includes several viruses than remain unassigned to any genus.

It was previously shown that entry into J cells mediated by nectin-GPI and nectin-EGFR becomes sensitive to bafilomycin A and is therefore similar to entry into J cells expressing both nectin1 and αVβ3-integrin (10). In this manner, ongoing infections may serve as adjuvants for subsequent infections by inducing costimulatory molecules and receptors that enhance APC function and recruitment. PML (red) is recruited to ICP4 associated foci that are distinct from those of 53BP1. We also reconstructed a tree from the sag-only alignment to evaluate the placement of sequences omitted from the main tree, and a pol-only tree to assess the relationship of sag-containing ERVs to closely related non-sag ERV. (a) A phylogeny of gag, pro, pol, env and sag genes. Clade-A shows the rodent ERVs, and RHVP, while clade-B is the predominantly primate clade that also contains AtHV3 and AtHV2. The rooting was determined by reconstructing a tree from an alignment of pol from sequences included in this tree as well as 11 outgroup sequences.

The scale bar shown represents the number of amino-acid substitutions per site and posterior probability values above 90% are indicated at each node. (b) WebLogo of a region of the alignment used to reconstruct the tree in a, including the underlying codons on the first line. The region shown is the crucial TCR activation motif. The subsequent lines show the codons for the same region using only clade-A or clade-B sequences, including a larger taxon set compared with the representative subset used for the tree. (c) Results of the Branch-Site test of positive selection, assessing the likelihood of positive selection at individual sites of the branch leading to SaHV2/AtHV3. The WebLogo was generated using the expanded clade-A alignment. Of these, 20 represent truncations or point mutants of miR-H2 through miR-H6, while 7 appear to represent random HSV-1 RNA breakdown products (Suppl.

The concatenated alignment revealed that there are two major lineages of sag—one belonging to retroviruses that primarily infect rodents (designated ‘clade-A’), and a second mainly primate-infecting ‘clade-B’ (Fig. The remainder of this work will elaborate a novel hypothesis , namely that a primordial herpes virus, rather than a transib or similar DNA transposon was the source of the primordial RAG-1 protein, and that understanding the biology of herpes virus replication is therefore essential to understanding the origins of the acquired immune system. Importantly, WT and R443I muSOX both degrade linear DNA with very similar kinetics (Figure 1E). We identified three independent lineages in the genome of Callithrix jacchus and Aotus nancymaae, which are not orthologous, despite the hosts having only recently speciated (~20 million years ago (mya)). Together, they form a robust monophyletic group that is consistent with a single-capture event of sag from a new world monkey retrovirus to the ancestor of the two new world monkey herpesviruses. Images reconstructed from electron micrographs of virions frozen in ice in the absence of stain, a technique by which morphology is better preserved, show that the core consists of the DNA packed at high density in liquid crystalline form, probably as a spool lacking a spindle (Booy et al., 1991; Zhou et al., 1999). Instead, RHVP sag is more closely related to rodent retroviruses, resolving as a sister taxon to J.

Convergent capture of retroviral superantigens by mammalian herpesviruses : Nature Communications : Nature Publishing Group
jaculus and P. Optionally, and preferably, the subject is first tested for the presence of a herpes virus, such as HSV-II, and if positive, the other steps are undertaken. Both herpesvirus sag lineages resolve within a clade of retroviruses to the exclusion of other retroviral sequences with high posterior probability, indicating that the direction of transfer was from retrovirus to herpesvirus. undetectable levels of infectious virus in homogenates of eyes or trigeminal ganglia (TG) at 35 or 70 days p.i. Cell proliferation was determined by seeding cells in six-well plates at a concentration of 2.5 × 104 cells/well. Most were found as ‘solo-LTRs’, which are the result of recombination of LTRs that leads to the deletion of the rest of the ERV. Relative, quantitative real-time RT-PCR was performed to quantify ICP0, gE, gD, and vhs mRNA levels over time using 18S rRNA internal controls as follows.

Briefly, a HindIII-PstI restriction fragment from pgC-mut (described below) was cotransfected into Vero cells with infectious KOS1.1 DNA. Notably, DNA binding proteins of the EBV replication complex and related proteins of other herpes viruses are highly antigenic proteins [39], [40] composing the so called “early antigen.” Expression of a herpes-like protein or proteins with regulated expression in the lymphocytes of an organism could immediately provide a potential selective advantage to the individual through stimulation of pathogen specific pattern receptors of the innate immune system such as toll receptors present in the sea urchin and similar invertebrates. In contrast, pre-treatment with Ad-ICP0 allowed HSV-1 0−GFP to express high levels of ICP0−GFP fluorescence (). It has been suggested that after the divergence into the salamander virus and frog virus lineages a subsequent host specific evolution could have occurred that would have limited cross transmission between both hosts, at least in laboratory infections [47]. In J-nectin1 cells, EIPA did not exert any inhibition, irrespective of the presence or absence of integrin. Second, TCRs may use two very different binding modes to recognize two unrelated ligands. These systems increase overall flexibility and allow straightforward triple label experiments.

sag-like BLAST hits (Fig. 2) that indicate a history of intragenomic proliferation. As in the case of the multilineage sags of mouse, rat and kangaroo rat, there are two distinct lineages in C. jacchus as well as a lineage in A. nancymaae (Fig. 4). (a) The Bayesian phylogenetic tree shown was reconstructed using only the sag partition of the alignment.

Albeit with lower support for deep nodes, the sag-only tree recovers the distinction between predominantly rodent- and primate-infecting viruses. The tree reveals that there are three independent lineages of rat ERVs, four lineages of mouse and two separate kangaroo rat clades. The scale bar shown represents amino-acid substitutions per site, and posterior probability values above 90% are indicated at each node. (b) The Bayesian phylogenetic tree shown was reconstructed using only the reverse transcriptase and integrase (RT and INT) partition of the alignment. Both clades A and B are observed as with the Fig. Cis-acting regulatory sequences immediately adjacent to the herpes DBP such as the BALF-2 protein of EBV, the only DBP for which experimental data is also available [48] have sequences resembling response elements for cellular factors such as AP-1 and also elements resembling other sites recognized by host transcription factors such as cAMP (cyclic AMP) and SP1 and AP-1 (Figure 5). The mean and standard deviation of the normalized mRNA/18S ratio is plotted for at least four independent experiments.

Starred tips indicate a pol of an ERV also included in the phylogenetic analysis of a concatenated alignment of gag, pol, env and sag. The scale bar shown represents amino-acid substitutions per site, and posterior probability values above 90% are indicated at each node. The pol tree (Fig. 4) allows us to reliably identify the phylogenetic placement of sag-containing ERVs identified in this study with respect to other retroviruses. It was possible to include ERVs that were identified as being closely related to MMTV, but did not contain recognizable sag according to our similarity cutoff. An example of such a compound is probenecid (4-(dipropylsulfamoyl)benzoic acid). The inclusion of known outgroup sequences in this tree confirms the distinction between clades A and B from which the separate herpesvirus sag acquisitions originate.

Reiterated DNA sequences in the RL region are denoted … TPA-induced and uninduced controls were prepared by incubating 106 TPA-induced and uninduced BCBL-1 cells in drug-free medium. 4), except for a lineage of Tarsius syrichta ERVs, which resolve at the root of the main phylogeny. Aliquots of labeled cell lysates were also analyzed by scintillation counting to determine total protein production and accumulation during each labeling period. This was done using the QuikChange site-directed mutagenesis system (Stratagene). A putative herpes virus recombinase pR1 homologous to RAG-1 with additional N-terminal amino acid sequences in both proteins [17] would also provide amino terminal protein sequences present in RAG-1, and bind to a primordial RAG-2 protein (pR2) co-expressed in somatic cells prior to the origins of the acquired immune system in the sea urchin. In contrast, when protein synthesis was inhibited for the first 6 hours of infection with cycloheximide, then both ICP0−GFP and ICP0+GFP fluorescent proteins accumulated to high levels by 6 hours post-release from the cycloheximide block ().

These data indicated that both combinations of genomic segments of SJNNV and RGNNV genotypes are successful and allow the resultant reassortant strains to produce disease. The raft-located CD4 interacts with and induces conformational changes to virion gp120. Cross-reactivity can alter the normal immunodominance hierarchy. Therefore it seems likely that similar events occur in infections by other viruses with similar DNA genomes and modes of nuclear entry, notably other members of the herpesvirus family. Both trees are shown on the same axis and have been pruned down to the taxa relevant to this study. Each of the hosts for which sag-containing ERVs could be dated through LTR divergence is marked by stars that indicate the age estimate obtained using different neutral rates of evolution. Only the highest and lowest values for R.

norvegicus are shown for clarity (denoted by an asterisk). The herpesvirus branches are drawn alongside the relevant host branch, to evaluate whether their respective divergence times are consistent with the null hypothesis of co-speciation. Yellow bars indicate the upper and lower bound age estimates. We cannot confidently reject a scenario of co-speciation for RHVP and MuHV4 since there is a known increase in the rate of evolution along this herpesvirus lineage, and the divergence date estimates are close to those of their hosts. The split between SaHV2 and AtHV3, however, is far younger than that of their hosts with no discernable rate increase. This discrepancy is consistent with cross-species transmission of the herpesviruses rather than co-speciation. The divergence date of the two monkey viruses therefore is a minimum bound for the acquisition of sag.

This date (~10 mya) is also similar to the integration date estimate of a closely related sag in the C. jacchus genome (~14 mya), which supports the existence of exogenous retroviruses circulating in new world monkeys, with a maximum bound at the root of the host clade ~25 mya. As discussed in the text, primordial RAG-2 protein (denoted pR2) may initially have blocked the recombinase functions of pR1 but exposed immunologic determinants essential to herpes virus immunity since the DBP are a major herpes virus antigen. We therefore reconstructed a time-calibrated phylogeny of rhadinoviruses using a relaxed log-normal molecular clock implemented in BEAST. We calibrated at previously identified co-diverging nodes (using host divergence dates23) that are outside the clades of interest, therefore enabling us to date the sag-containing herpesvirus splits without assuming co-divergence a priori. This revealed that the SaHV2-AtHV3 split occurred ~10.6 mya (95% HPD: 8.4–13), while that of RHVP and its closest relative MuHV4 diverged~37.5 mya (95% highest posterior density (HPD): 32.7–42.6; Fig. 5).

The results from the phylogenetic analysis support the hypothesis that sag was co-opted by herpesviruses from retroviruses. If this is true, the retroviral donors must have possessed functional sag, and so we sought evidence for a history of purifying selection in the sag-like sequences that we identified. The compounds are precursors of acyclovir, penciclovir, ganciclovir or others, and are administered in doses equivalent in activity to a standard famciclovir (FAMVIR-Novartis, generic version from Teva Pharmaceuticals) as taught in the provisional and PCT applications mentioned above. We estimated average dN/dS along branches in the tree (the so-called branch models) by allowing every lineage to have its own ω (free-ratio test) or fixing the tree to a single estimate (one-ratio test). ). In contrast, BDCRB, 1263W94, and 175X were all inactive against the three alphaviruses tested. This means that their endogenization was relatively recent (which is consistent with the results of the LTR dating) and these sequences were likely functional in the past.

Overall, the proteins that appeared most stable in WT- and UL49R-infected cells were also most stable in UL49−-infected cells. In others, the probe was the linearized pEcoRI-BamHI-1-1 plasmid (18). Conserved D and E residues in ICP-8, other herpes DBP and RAG proteins (defined in ) are in proximity and thus capable of contacting a bound double stranded nucleic acid based upon their positions in the partial ICP-8 structure (). Regardless of their different genotypes, 2.5 pfu per cell of each HSV-1 VP16+ virus yielded high and equivalent levels of ICP0 mRNA in cycloheximide-treated cultures (). The use of phylogenetic tools has provided considerable genetic evidence indicating that rainbow trout pathogenic VHSV emerged from a genotype I-type marine ancestor [98,101-103].

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Convergent capture of retroviral superantigens by mammalian herpesviruses : Nature Communications : Nature Publishing Group

Convergent capture of retroviral superantigens by mammalian herpesviruses : Nature Communications : Nature Publishing Group

The etiology of many autoimmune diseases such as systemic lupus erythematosus, progressive systemic sclerosis (scleroderma), and psoriasis vulgaris remains unclear. HSV expression vectors overcome these disadvantages, but, because they persist in target cells as nonreplicative episomes, they are not retained in all the progeny of dividing cells. Restriction of HSV-1 replication and progeny production is important to prevent viral spread, but the cellular mechanisms that inhibit the DNA virus in macrophages are unknown. Peripheral blood for FACS immunophenotyping, isolation of peripheral mononuclear cells (PMNC), and serum ELISA assays for IL-12 and soluble Fas ligand (sFasL) was collected daily during the first 4 post-operative weeks. We further show through a timescaled phylogenetic analysis that a cross-species transmission of monkey herpesviruses occurred after the acquisition of sag. These results reveal that a diverse range of ancient sag-containing retroviruses independently donated sag twice from two separate lineages that are distinct from MMTV. These vectors have wide applications for the genetic modification of many cell types.

Here, we demonstrate that SAMHD1 restricts replication of the HSV-1 DNA genome in differentiated macrophage cell lines. Serum ELISA revealed significantly increased sFasL and IL-12 levels in the gene therapy group compared with controls. (b) Maximum-likelihood phylogenies of host IL-17 and CD59 from primates including a viral homologue in SaHV2. SaHV2 independently acquired these genes after diverging from AtHV3. The gene for CD59 in particular, appears to have been captured from the squirrel monkeys, which are the current host of the virus. Our results suggest that SAMHD1 functions broadly to inhibit replication of DNA viruses in nondividing macrophages. Gene Therapy (2000) 7, 1853-1858.

The results of 1,000 nonparametric bootstrap replicates are indicated at each node. The histogram shows the number of tBLASTn hits per mammalian genome to any of the four viral sags. A conserved 100 amino-acid region of the sag sequences of SaHV2, AtHV3, RHVP and MMTV was used as a query. The mouse, rat, Chinese hamster and hedgehog data represent multiple individual genomes. Patients were randomly assigned to the gene therapy or control arms (Table 1). (a) A phylogeny of gag, pro, pol, env and sag genes. Clade-A shows the rodent ERVs, and RHVP, while clade-B is the predominantly primate clade that also contains AtHV3 and AtHV2.

Convergent capture of retroviral superantigens by mammalian herpesviruses : Nature Communications : Nature Publishing Group
The rooting was determined by reconstructing a tree from an alignment of pol from sequences included in this tree as well as 11 outgroup sequences. The scale bar shown represents the number of amino-acid substitutions per site and posterior probability values above 90% are indicated at each node. Analysis of primary tumors demonstrated minor to moderate intratumoral T lymphocytic infiltrates with no significant B cell accumulation, and perivascular populations of activated microglial cells (Table 2). The region shown is the crucial TCR activation motif. The subsequent lines show the codons for the same region using only clade-A or clade-B sequences, including a larger taxon set compared with the representative subset used for the tree. (c) Results of the Branch-Site test of positive selection, assessing the likelihood of positive selection at individual sites of the branch leading to SaHV2/AtHV3. The WebLogo was generated using the expanded clade-A alignment.

Numbers of T cells (CD3+) and CD4+ or CD8+ T cell subsets (helper and cytotoxic lymphocytes, respectively) did not display a significant increase during GCV therapy (days 14-27), and did not differ significantly in either group of patients (Figure 2a-c). (a) The Bayesian phylogenetic tree shown was reconstructed using only the sag partition of the alignment. Albeit with lower support for deep nodes, the sag-only tree recovers the distinction between predominantly rodent- and primate-infecting viruses. The tree reveals that there are three independent lineages of rat ERVs, four lineages of mouse and two separate kangaroo rat clades. The scale bar shown represents amino-acid substitutions per site, and posterior probability values above 90% are indicated at each node. In the control group, serum sFasL levels were below the detection limit of the assay (5 pg/ml). Both clades A and B are observed as with the Fig.

3 and sag-only tree in the upper panel. In addition, the RT–INT tree shows that clade-A is closely related to a large and diverse group of sag-less mammalian ERVs. Starred tips indicate a pol of an ERV also included in the phylogenetic analysis of a concatenated alignment of gag, pol, env and sag. The reactivity of control patients’ PMNCs against VPC and autologous tumor was significantly lower compared with those values in gene therapy patients (Figure 4). The figure depicts time-calibrated phylogenies of Herpesviruses (dark grey) and mammalian hosts (light grey). Both trees are shown on the same axis and have been pruned down to the taxa relevant to this study. Each of the hosts for which sag-containing ERVs could be dated through LTR divergence is marked by stars that indicate the age estimate obtained using different neutral rates of evolution.

Only the highest and lowest values for R. During HSV-tk/GCV pharmacogene therapy, no radiation therapy was applied. The herpesvirus branches are drawn alongside the relevant host branch, to evaluate whether their respective divergence times are consistent with the null hypothesis of co-speciation. Yellow bars indicate the upper and lower bound age estimates. We cannot confidently reject a scenario of co-speciation for RHVP and MuHV4 since there is a known increase in the rate of evolution along this herpesvirus lineage, and the divergence date estimates are close to those of their hosts. The split between SaHV2 and AtHV3, however, is far younger than that of their hosts with no discernable rate increase. In conclusion, our findings, as well as previous reports15,16,17,18 on an immunological component of HSV-tk/GCV gene therapy may reveal an additional effect of pharmacogene therapy: destruction of tumor cells by bioactivated drugs provides tumor antigen release without suppression of antitumor immune response, as with conventional chemotherapy or radiation.

The divergence date of the two monkey viruses therefore is a minimum bound for the acquisition of sag. This date (~10 mya) is also similar to the integration date estimate of a closely related sag in the C. jacchus genome (~14 mya), which supports the existence of exogenous retroviruses circulating in new world monkeys, with a maximum bound at the root of the host clade ~25 mya.

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