Simian Foamy Virus Type Research Articles

Identification of the simian foamy virus transcriptional transactivator gene (taf)

Simian foamy virus type 1 (SFV-1), a member of spumavirus subfamily of retroviruses, encodes a transcriptional transactivator that functions to strongly augment gene expression directed by the viral long terminal repeat (LTR). The objective of this study was to identify the viral gene responsible for transactivation. Nucleotide sequences between the env gene and the LTR of SFV-1 were determined. The predicted amino acid sequence revealed two large open reading frames (ORFs), designated ORF-1 (311 amino acids) and ORF-2 (422 amino acids). In the corresponding region of the human foamy virus, three ORFs (bel-1, bel-2, and bel-3) have been identified (R. M. Flugel, A. Rethwilm, B. Maurer, and G. Darai, EMBO J. 6:2077-2084, 1987). Pairwise comparisons of the ORF-1 and ORF-2 with bel-1 and bel-2 show small clusters of homology; less than 39% overall homology of conserved amino acids is observed. A counterpart for human foamy virus bel-3 is not present in the SFV-1 sequence. Three species of viral RNA have been identified in cells infected with SFV-1; an 11.5-kb RNA representing full-length transcripts, a 6.5-kb RNA representing the env message, and a 2.8-kb RNA from the ORF region. Analysis of a cDNA clone encoding the ORF region of SFV-1 reveals that the 2.8-kb message is generated by complex splicing events involving the 3' end of the env gene. In transient expression assays in cell lines representing several species. ORF-1 was shown to be necessary and sufficient for transactivating viral gene expression directed by the SFV-1 LTR. The target for transactivation is located in the U3 domain of the LTR, upstream from position - 125 (+ 1 represents the transcription initiation site). We propose that OFF-1 of SFV-1 be designated the transcriptional transactivator of foamy virus (taf).

Sequence analysis of the simian foamy virus type 1 genome.

We have cloned the simian foamy virus type 1 genome (SFV1) and determined its nucleotide sequence. Analysis of this genome reveals, in addition to the usual genes encoding retroviral capsid, reverse transcriptase, and envelope protein (respectively, gag, pol, and env), two open reading frames (ORFs) between env and the long terminal repeat with partial homology to the human foamy virus (HFV) bel1 and bel2 genes. The first ORF could code for a polypeptide of 312 amino acids (aa) showing 40% homology with the HFV bel1 putative gene product. A more detailed analysis showed that the protein encoded by this ORF would have features characteristic of known trans-activating proteins. The second ORF could code for a polypeptide of 403 aa showing 38% homology with the putative HFV bel2 gene product. Moreover, the 5' extremity of the RNA genome can be folded into a secondary structure identical to the Tat-response element of human immunodeficiency viruses. A phylogenetic tree of retroviruses, including SFV1 and HFV, was constructed. It showed at the molecular level that Spumavirinae, previously classified on the basis of their morphology and their biological properties, constitute a separate group. The homology between SFV1 and HFV reaches 89% in the reverse transcriptase domain of the pol gene. but is much smaller in other parts of the genome.

Simian foamy virus type 1 is a retrovirus which encodes a transcriptional transactivator

Simian foamy viruses, members of the spumavirus subfamily of retroviruses, are found in a variety of nonhuman primates and, as yet, remain to be characterized with respect to genetic structure and regulation of viral gene expression. The genome of simian foamy virus type 1 (SFV-1), an isolate from rhesus macaques, has been molecularly cloned, and the role of the viral long terminal repeat (LTR) in transcriptional control has been investigated. The SFV-1 LTR is 1,621 base pairs long, and sequence comparisons with human foamy virus revealed a pattern of clustered homology. A cap site in the LTR was identified by analysis of SFV-1 transcripts in infected cells. Transient expression assays in cell lines representing several species and different cell types showed that the SFV-1 LTR has low basal activity in uninfected cells, whereas LTR-directed expression is greatly increased in cells infected with SFV-1. This transactivation is mediated by a mechanism involving increases in steady-state levels of viral transcripts. Thus, the SFV-1 genome encodes a transactivator that functions on the LTR at the transcriptional level.

Relationship of the env genes and the endonuclease domain of the pol genes of simian foamy virus type 1 and human foamy virus

We have molecularly cloned and sequenced a portion of the simian foamy virus type 1 (SFV-1); open reading frames representing the endonuclease domain of the polymerase (pol) and the envelope (env) genes were identified by comparison with the human foamy virus (HFV). Unlike the HFV genomic organization, the SFV-1 pol gene overlaps the env gene; thus, the open reading frames reported for HFV between pol and env is not present in SFV-1. Comparisons of predicted amino acid sequences of HFV and SFV-1 reveal that the endonuclease domains of the pol genes are about 84% related. The region predicted to encode the SFV-1 extracellular env domain is 569 codons; SFV-1 and HFV have 64% amino acid similarity in this env domain. The predicted hydrophobic transmembrane env proteins of both HFV and SFV-1 show about 73% similarity. A total of 16 potential glycosylation sites are found in SFV-1 env, and 15 are found in HFV; 11 are shared. SFV-1 has 25 cysteine residues, and HFV has 23 residues; all 23 cysteine residues of HFV are conserved in SFV-1. This sequence analysis reveals that the human and simian foamy viruses are highly related.

Effects of human recombinant alpha and gamma and of highly purified natural beta interferons on simian Spumavirinae prototype (simian foamy virus 1) multiplication in human cells

The present study demonstrates the inhibitory effect of human recombinant interferons (r-Hu-IFN) alpha and gamma, and that of highly purified natural human interferon beta on the replication of simian foamy virus type 1 (SFV1) in human AV3-cell cultures. All IFN led to strong inhibition of the SFV1 cytopathic effect. Electron microscopy showed a 70 to 95 % decrease in viral particles. Significant inhibition of virus-associated reverse transcriptase activity was found in supernatant fluids of infected IFN-treated cultures. Metabolic labelling of the virus confirmed the inhibition of extracellular release of SFV1. PAGE analysis of immunoprecipitates indicated a reduction in viral-specific protein bands. Altogether, these results indicate that the mechanism of inhibition of Spumavirinae infection by interferon differs from that described for the other Retroviridae, and particularly for types B, C and D viruses. Our data is of therapeutic interest since Spumavirinae have been linked to pathological processes such as de Quervain thyroiditis.

Heterogeneity of primate foamy virus genomes

DNA of the T-lymphotropic simian foamy virus (SFV) LK-3 was cloned in a plasmid vector and used as a probe in comparative DNA:DNA hybridization studies with primate foamy viruses (SFV serotypes 1, 2, 3, 5, 6, 7, 8, fresh SFV isolates from 12 African green monkeys and one rhesus monkey, as well as human syncytium forming virus). All freshly isolated viruses seemed to be variants of SFV types 2 and 3. Especially LK-3 appeared to be closely related to SFV type 3. No hybridization of the LK-3 clone was observed to DNA sequences of human immunodeficiency virus (HIV-1).

Characterization of a Human Lymphoblastoid Cell Line Permanently Modified by Simian Foamy Virus Type 10

Simian Spumavirinae serotype, SFV10, of a Papio cynocephalus baboon, was used to infect a human lymphoblastoid cell line, LV2. Permanent growth and morphological alterations of infected cells occurred, even though no viral particles were detected. Evidence for the presence of viral genomes in the modified cell line is provided indirectly from immunological studies and induction experiments followed by coculture procedures. The permanently modified cell line obtained (LV2-FB10) is an interesting model for the investigation of the possible integration of foamy viruses into the host genomes.

Cell transfection with DNA from simian foamy virus type 1 producing cultures generates equivalent infectious virions

Summary The transfection of canine and murine cells with DNA from simian foamy virus type 1 cell cultures led to the appearance of a cytopathic effect (CPE) similar to that induced by the original SFV1. High reverse transcriptase activity was present in supernatants of transfected cultures. Inoculation of these supernatants into permissive cells induced a syncytial CPE and high RNA-dependent DNA polymerase activity. These two effects were specifically inhibited by immune serum against SFV1. By electron microscopy, inoculated cultures showed the presence of virions homologous to those of SFV1.

Purification and characterization of simian foamy virus type I structural core polypeptides.

The present study concerns the purification and partial characterization of simian foamy virus type 1 (SFV 1) structural core polypeptides. The obtention of SFV 1 cores separated from envelope components after viral disrupture was verified by electron microscopy (EM), density gradient, and polyacrylamide gel electrophoresis (PAGE). Multistep purification by column chromatography, verified by PAGE, enabled us to separate the structural core polypeptides from the 80,000 molecular weight reverse transcriptase. Two species of structural core polypeptides were identified with apparent molecular weights of 51 and 15 kd. By affinity chromatography on a double-stranded DNA-cellulose column, the main internal protein, p51, was shown to be composed of a major 30 kd protein and a minor 19 kd polypeptide, which binds to double stranded DNA. The p15 internal protein was shown to have a ribonucleotide binding nature.

Appearance of Epstein-Barr virus early antigen (EA) in Raji cells infected with retroviruses

Summary Infection of Raji cells (human lymphoblastoid cells carrying the Epstein-Barr (EB) virus) with xenotropic retroviruses of different taxonomic subgroups activated the EB early antigens, which are normally not expressed in these cells. The viruses used were type C feline, baboon and Mus molossinus mouse endogenous viruses, type C gibbon ape virus, type D Mason Pfizer monkey virus and simian foamy virus type A. Syncytia formation was consistently observed in infected cultures. The significance of the antigen induction and possible mechanisms of action are discussed.

Purification and Characterization of the Major Envelope Glycoprotein of Simian Foamy Virus Type 1

Simian foamy virus type 1 (SFV-1), the prototype of the Spumavirinae, was subjected to disruption and serial purification procedures. Separation of SFV-1 envelope components from viral cores was verified by electron microscopy, density gradient centrifugation and polyacrylamide gel electrophoresis. After affinity chromatography of the envelope polypeptides on a concanavalin A-Sepharose column, a highly purified 70 000 mol. wt. protein was recovered. Glycosylation of this gp70 was confirmed by glucosamine labelling. Immunological studies with anti-SFV antisera confirmed the type-specificity of this envelope gp70.

Characterization of RNase H activity associated with reverse transcriptase in simian foamy virus type 1

Spumavirinae or foamy viruses have been shown to have a characteristic RNA-dependent DNA polymerase activity. We demonstrate here the existence of an RNase H activity that copurifies with the 81-kilodalton monomeric polypeptide, which carries the RNA-dependent DNA polymerase activity of simian foamy virus type 1. RNase H degrades RNA hybrid substrates; however, it does not solubilize single-stranded RNAs. Inactivation assays with heat, high levels of bivalent cations, ethidium bromide, and sodium fluoride suggest that the RNase H catalytic site could be topologically independent from the DNA polymerase catalytic site.

Reverse transcriptase from simian foamy virus serotype 1: purification and characterization.

Chromatography on heparin-Sepharose, known for its affinity for nucleotide-binding polypeptides, was used to purify the viral RNA-dependent DNA polymerase (reverse transcriptase) from the core polypeptides of simian foamy virus type 1. This procedure allowed the recovery of highly purified enzyme with a high specific activity. The average molecular weight of this monomeric enzyme is 81,000 and is thus comparable to that found for other known primate retroviruses. Reverse transcriptase activity of simian foamy virus type 1 requires a ribonucleotide template as a primer or otherwise a DNA with 3'-OH ends. Other optimal conditions of activity are reviewed. Heat inactivation studies led to the concept of an enzyme with two loci, one specific for the substrate and the other for the template-primer.

Persistent asymptomatic infection of the laboratory mouse by simian foamy virus type 6: a new model of retrovirus latency.

Simian foamy virus (SFV) type 6, originally isolated from the kidney of a kuru-inoculated chimpanzee, has been adapted to produce an asymptomatic infection in Swiss-Webster white mice, with virus detected in the kidney and spleen for up to 10 months after intra-peritoneal inoculation, and the presence in some animals of complement-fixing, but not neutralizing, serum antibody. This first successful experimental infection of the laboratory mouse by a representative of the foamy virus group has special interest as a convenient model in which to study pathogenesis and viral latency, particularly in view of the isolation and recent characterization of a foamy virus from human tissue which is virtually indistinguishable from the SFV type 6 used in the present study.

Biochemical characterization of simian foamy virus type i.

Simian syncitium-forming ("foamy") virus type I (SFV1) was characterized biochemically. RNA was extracted from purified virus either with 0.1 per cent SDS or by the standard phenol-chloroform method. By both techniques a main component of 65-70S was found. Denaturation of the 65-70S RNA by heat resulted in a shift of the sedimentation coefficient mainly to a 30-35S component. Electrophoresis on a composite polyacrylamide gel demonstrated the existence of three minor RNA's: 8S, 5S and 4S respectively. PAGE-SDS analysis of disrupted purified virions enabled the separate migration of five viral proteins and the identification of two main proteins: a 30 kd polypeptide and a 70 kd polypeptide.

Characterization of a simian foamy virus isolated from a spontaneous primate lymphoma.

Examination of tissues from a stump-tailed monkey (Macaca arctoides) with a spontaneous lymphoid malignancy by light and electron microscopy confirmed the presence of a diffuse lymphoma of intermediate cell type. Membranes showed positivity with anti-Ia-like antisera and negativity with all other markers. In vitro cocultivation of affected tissues resulted in the isolation of an agent found to have properties idenitical to those described for simian foamy virus type 1. There was no evidence for the presence of oncoviruses.

Human Foamy Virus: Further Characterization, Seroepidemiology, and Relationship to Chimpanzee Foamy Viruses

A foamy virus present in human nasopharyngeal carcinoma tissue was studied for a number of biological properties, including range of cellular susceptibility, growth curve, evolution of cytopathic effect in relation to cellular fusion and intracellular viral distribution, reverse transcriptase activity, and buoyant density. The virus was also studied immunologically and found to be closely related to the chimpanzee foamy viruses, particularly simian foamy virus type 6, with which it shares common antigens in complement-fixing, fluorescent, and neutralizing antibody tests. In view of this close immunological relationship and the failure to find antibody to the human isolate in sera from more than 250 humans, including 50 patients with nasopharyngeal carcinoma and Burkitt's lymphoma, it is suggested that the isolate is not a human representative of the foamy virus group but rather a variant strain of chimpanzee foamy virus.

A Haemagglutination Assay for the Primate Syncytium-forming (Foamy) Viruses

Preparations of highly concentrated simian foamy virus type 1 (SFV1)agglutinate guinea pig red blood cells. The agglutination occurs both at 22 degrees and 37 degrees C. It is inhibited by specific antisera against SFV1 but also by antisera prepared against several other types of simian foamy viruses.

Pathogenesis of simian foamy virus infection in natural and experimental hosts

Inoculation of simian foamy virus type 1 into New Zealand white rabits resulted in an infection which was very similar to that observed in naturally infected nonhuman primates. Both intraperitoneal and intranasal inculations were found to be efficient procedures for the establishment of the infection in rabbits. Infection by the nasal route was found to be the best method, whereas no infection could be established by feeding virus in the drinking water. Once infection was established, virus persisted in the tissues and organs for as long as 264 days after inoculation, during which time the animals maintained significant levels of neutralizing antibody. Infectious virus was recovered from spleen, liver, lung, salivary gland, kidney, and, to a lesser extent, the brain. Virus was isolated from the blood only during early infection and never from the urine. A comparison of the distribution of foamy virus in naturally infected monkeys and baboon with experimentally infected rabbits showed that both groups harbored infectious virus in the same internal tissues and organs. Recovery of infectious virus from both groups of animals was accomplished by cultivation and/or co-cultivation of infected cells onto Vero cells.