Reticuloendotheliosis type C and primate type D oncoretroviruses are members of the same receptor interference group.

The reticuloendotheliosis viruses (REVs), originally isolated from avian species, constitute a group of retroviruses which are more closely related to mammalian retroviruses than to other avian retroviruses. The envelope glycoproteins of members of the REV group display a striking amino acid sequence identity with a group of primate oncoretroviruses which belong to a single receptor interference group and include all of the type D and some type C primate oncoretroviruses. Members of the REV group also have a broad host range which covers most avian cells and some mammalian cells, including those of simian and human origin. In view of this broad host range and the envelope sequence similarities, we investigated the cross-interference pattern between REV and primate virus groups to determine whether they utilized the same receptor. Superinfection experiments using a vector virus containing an Escherichia coli lacZ gene showed that reticuloendotheliosis and simian oncoretroviruses constitute a single receptor interference group on both human and canine cells and indicate that the viruses bind to the same receptor to initiate infection. These results suggest that this receptor binding specificity has been maintained over a wide range of retroviruses and may be responsible for the broad spread of these retroviruses between different orders of vertebrates.     

Spleen necrosis virus, an avian immunosuppressive retrovirus, shares a receptor with the type D simian retroviruses.

The reticuloendotheliosis viruses (REV) are a family of highly related retroviruses isolated from gallinaceous birds. On the basis of sequence comparison and overall genome organization, these viruses are more similar to the mammalian type C retroviruses than to the avian sarcoma/leukemia viruses. The envelope of a member of the REV family, spleen necrosis virus (SNV), is about 50% identical in amino acid sequence to the envelope of the type D simian retroviruses. Although SNV does not productively infect primate or murine cells, the receptor for SNV is present on a variety of human and murine cells. Moreover, interference assays show that the receptor for SNV is the same as the receptor for the type D simian retroviruses. We propose that adaptation of a mammalian type C virus to an avian host provided the REV progenitor.     

Endogenous bornavirus elements in mammalian genome

Approximately 8% of our genome is made up of endogenous retroviral elements. Endogenous retrovirus is a fossil record of ancient retrovirus infection and, therefore, gives important insights into the evolutional relationship between retroviruses and their hosts. On the other hand, until recently, it has been believed that no endogenous non-retroviral viruses exist in animal genomes. We lately discovered endogenous elements homologous to the nucleoprotein of bornaviruses, a negative-strand RNA virus, in the genomes of many mammalian species, including humans. We also demonstrated that mRNA of extant mammalian bornavirus, Borna disease virus, is reverse-transcribed and integrated into the host genome DNA. These findings provided novel insights not only into the interaction between RNA viruses and their hosts, but also into the mechanism underlying the gain of novelty in mammalian genomes. In this review, we will briefly summarize our recent knowledge about endogenous bornavirus elements and also introduce some recent discoveries regarding endogenous elements of non-retroviral viruses in vertebrate genomes.     

Endogenous viral elements in animal genomes

Integration into the nuclear genome of germ line cells can lead to vertical inheritance of retroviral genes as host alleles. For other viruses, germ line integration has only rarely been documented. Nonetheless, we identified endogenous viral elements (EVEs) derived from ten non-retroviral families by systematic in silico screening of animal genomes, including the first endogenous representatives of double-stranded RNA, reverse-transcribing DNA, and segmented RNA viruses, and the first endogenous DNA viruses in mammalian genomes. Phylogenetic and genomic analysis of EVEs across multiple host species revealed novel information about the origin and evolution of diverse virus groups. Furthermore, several of the elements identified here encode intact open reading frames or are expressed as mRNA. For one element in the primate lineage, we provide statistically robust evidence for exaptation. Our findings establish that genetic material derived from all known viral genome types and replication strategies can enter the animal germ line, greatly broadening the scope of paleovirological studies and indicating a more significant evolutionary role for gene flow from virus to animal genomes than has previously been recognized.     

Genome of Reticuloendotheliosis Virus: Characterization by Use of Cloned Proviral DNA

Reticuloendotheliosis virus is an avian type C retrovirus that is capable of transforming fibroblasts and hematopoietic cells both in vivo and in vitro. This virus is highly related to the three other members of the reticuloendotheliosis virus group, including spleen necrosis virus, but it is apparently unrelated to the avian leukosis-sarcoma virus family. Previous studies have shown that it consists of a replication-competent helper virus (designated REV-A) and a defective component (designated REV) that is responsible for transformation. In this study we used restriction endonuclease mapping and heteroduplex analysis to characterize the proviral DNAs of REV-A and REV. Both producer and nonproducer transformed chicken spleen cells were used as sources of REV proviral DNA; this genome was mapped in detail, and fragments of it were cloned in lambdagtWES.lambdaB. The infected canine thymus line Cf2Th(REV-A) was used as a source of REV-A proviral DNA. The restriction maps and heteroduplexes of the REV and REV-A genomes showed that (proceeding from 5' to 3') (i) REV contains a large fraction of the REV-A gag gene (assuming a gene order of gag-pol-env and gene sizes similar to those of other type C viruses), for the two genomes are very similar over a distance of 2.1 kilobases beginning at their 5' termini; (ii) most or all of REV-A pol is deleted in REV; (iii) REV contains a 1.1 kilobase segment derived from the 3' end of REV-A pol or the 5' end of env or both; (iv) this env region in REV is followed by a 1.9-kilobase segment which is unrelated to REV-A; and (v) the helper-unrelated segment of REV extends essentially all of the way to the beginning of the 3' long terminal repeat. Therefore, like avian myeloblastosis virus but unlike the other avian acute leukemia viruses and most mammalian and avian sarcoma viruses, REV appears to be an env gene recombinant. We also found that the REV-specific segment is derived from avian DNA, for a cloned REV fragment was able to hybridize with the DNA from an uninfected chicken. Therefore, like the other acute transforming viruses, REV appears to be the product of recombination between a replication-competent virus and host DNA. Two other defective genomes in virus-producing chicken cells were also cloned and characterized. One was very similar to REV in its presumptive gag and env segments, but instead of a host-derived insertion it contained additional env sequences. The second was similar (but not identical) to the first in its gag and env regions and appeared to contain an additional 1-kilobase inversion of REV-A sequences.     

Reticuloendotheliosis virus: detection of immunological relationship to mammalian type C retroviruses.

Reticuloendotheliosis virus (REV) p30 shares cross-reactive determinants and a common NH2-terminal tripeptide with mammalian type C viral p30's. An interspecies competition radioimmunoassay was developed, using iodinated REV p30 and a broadly reactive antiserum to mammalian virus p30's. The avian leukosis-sarcoma viruses and mammalian non-type C retroviruses did not compete in this assay. Previous data indicating that the REV group is not represented completely in normal avian cell DNA lead us to speculate that this may be the first example of interclass transmission, albeit in the remote past, among the Retroviridae.     

Infectious endogenous retroviruses in cats and emergence of recombinant viruses

Endogenous retroviruses (ERVs) comprise a significant percentage of the mammalian genome, and it is poorly understood whether they will remain as inactive genomes or emerge as infectious retroviruses. Although several types of ERVs are present in domestic cats, infectious ERVs have not been demonstrated. Here, we report a previously uncharacterized class of endogenous gammaretroviruses, termed ERV-DCs, that is present and hereditary in the domestic cat genome. We have characterized a subset of ERV-DC proviral clones, which are numbered according to their genomic insertions. One of these, ERV-DC10, located in the q12-q21 region on chromosome C1, is an infectious gammaretrovirus capable of infecting a broad range of cells, including human. Our studies indicate that ERV-DC10 entered the genome of domestic cats in the recent past and appeared to translocate to or reintegrate at a distinct locus as infectious ERV-DC18. Insertional polymorphism analysis revealed that 92 of 244 domestic cats had ERV-DC10 on a homozygous or heterozygous locus. ERV-DC-like sequences were found in primate and rodent genomes, suggesting that these ERVs, and recombinant viruses such as RD-114 and BaEV, originated from an ancestor of ERV-DC. We also found that a novel recombinant virus, feline leukemia virus subgroup D (FeLV-D), was generated by ERV-DC env transduction into feline leukemia virus in domestic cats. Our results indicate that ERV-DCs behave as donors and/or acceptors in the generation of infectious, recombinant viruses. The presence of such infectious endogenous retroviruses, which could be harmful or beneficial to the host, may affect veterinary medicine and public health.     

Endogenous Hepatitis C Virus Homolog Fragments in European Rabbit and Hare Genomes Replicate in Cell Culture

Endogenous retroviruses, non-retroviral RNA viruses and DNA viruses have been found in the mammalian genomes. The origin of Hepatitis C virus (HCV), the major cause of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma in humans, remains unclear since its discovery. Here we show that fragments homologous to HCV structural and non-structural (NS) proteins present in the European rabbit (Oryctolagus cuniculus) and hare (Lepus europaeus) genomes replicate in bovine cell cultures. The HCV genomic homolog fragments were demonstrated by RT-PCR, PCR, mass spectrometry, and replication in bovine cell cultures by immunofluorescence assay (IFA) and immunogold electron microscopy (IEM) using specific MAbs for HCV NS3, NS4A, and NS5 proteins. These findings may lead to novel research approaches on the HCV origin, genesis, evolution and diversity.     

Spontaneous heteromerization of gammaretrovirus envelope proteins: a possible novel mechanism of retrovirus restriction

The env gene of gammaretroviruses encodes a glycoprotein conserved among diverse retroviruses, except for the domains involved in receptor binding. Here we show that pairs of gammaretrovirus envelope proteins (from Friend virus and GALV or xenotropic viruses) assemble into heteromers when coexpressed. This assembly results in a strong inhibition of infectivity. An unrelated envelope protein does not assemble in heteromers with the gammaretrovirus glycoproteins tested and does not affect their infectivity, demonstrating the specificity of the mechanism we describe. We propose that the numerous copies of endogenous retroviral env genes conserved within mammalian genomes act as restriction factors against infectious retroviruses.     

An Endogenous Foamy-like Viral Element in the Coelacanth Genome

Little is known about the origin and long-term evolutionary mode of retroviruses. Retroviruses can integrate into their hosts'' genomes, providing a molecular fossil record for studying their deep history. Here we report the discovery of an endogenous foamy virus-like element, which we designate ‘coelacanth endogenous foamy-like virus’ (CoeEFV), within the genome of the coelacanth (Latimeria chalumnae). Phylogenetic analyses place CoeEFV basal to all known foamy viruses, strongly suggesting an ancient ocean origin of this major retroviral lineage, which had previously been known to infect only land mammals. The discovery of CoeEFV reveals the presence of foamy-like viruses in species outside the Mammalia. We show that foamy-like viruses have likely codiverged with their vertebrate hosts for more than 407 million years and underwent an evolutionary transition from water to land with their vertebrate hosts. These findings suggest an ancient marine origin of retroviruses and have important implications in understanding foamy virus biology.     

Specific antigenic relationships between the RNA-dependent DNA polymerases of avian reticuloendotheliosis viruses and mammalian type C retroviruses

Immunoglobulin G directed against the DNA polymerase of Rauscher murine leukemia virus (R-MuLV) could bind to 125I-labeled DNA polymerase of spleen necrosis virus (SNV), a member of the reticuloendotheliosis virus (REV) species. Competition radioimmunoassays showed the specificity of this cross-reaction. The antigenic determinants common to SNV and R-MuLV DNA polymerases were shared completely by the DNA polymerases of Gross MuLV, Moloney MuLV, RD 114 virus, REV-T, and duck infectious anemia virus. Baboon endogenous virus and chicken syncytial virus competed partially for antibodies directed against the common antigenic determinants of SNV and R-MuLV DNA polymerases. DNA polymerases of avian leukosis viruses, pheasant viruses, and mammalian type B and D retroviruses and particles with RNA-dependent DNA polymerase activity from the allantoic fluid of normal chicken eggs and from the medium of a goose cell culture did not compete for the antibodies directed against the common antigenic determinants of SNV and R-MuLV DNA polymerases. We also present data about a factor in normal mammalian immunoglobulin G that specifically inhibits the DNA polymerases of REV and mammalian type C retrovirus DNA polymerases.     

ART-CH, a new chicken retroviruslike element.

A 3' region of a previously unknown retroviruslike element named ART-CH (avian retrotransposon from chicken genome) was obtained in the course of polymerase chain reaction-mediated cloning of avian leukosis virus long terminal repeats (LTRs) from DNAs of infected chicken cells. About 50 copies of ART-CH are present in the genome of chickens of different breeds. ART-CH is not found in DNA of quails, ducks, turkeys, or several other birds tested. The ART-CH element is about 3 kb in size, including 388 bp LTRs. The major class of ART-CH-specific RNA, also 3 kb in size, is detected in various organs of chickens. An ART-CH polypurine tract, a tRNA(Trp)-binding site, regions around the TATA box and polyadenylation signal, and the beginning of the putative gag gene strongly resemble the corresponding regions of avian leukosis viruses and EAV, the two described classes of chicken retroviruses. An open reading frame capable of encoding a polypeptide with a putative transmembrane domain is located upstream of the right ART-CH LTR. This sequence, as well as the U3 and U5 regions of the ART-CH LTR, has no obvious similarities with the corresponding parts of other known vertebrate retroviruses and retrotransposons. A short sequence upstream of the right LTR of ART-CH is very similar to sequences which flank the 3' ends of the oncogenes v-src, v-myc, v-fps, and v-crk in four different recombinant avian retroviruses and which are absent from the genomes of other studied avian retroviruses. Thus, ART-CH is a new endogenous chicken provirus that may participate in the formation of recombinant oncogenic retroviruses.     

Evolution and distribution of class II-related endogenous retroviruses

Endogenous retroviruses (ERVs) are widespread in vertebrate genomes and have been loosely grouped into "classes" on the basis of their phylogenetic relatedness to the established genera of exogenous retroviruses. Four of these genera-the lentiviruses, alpharetroviruses, betaretroviruses, and deltaretroviruses-form a well-supported clade in retroviral phylogenies, and ERVs that group with these genera have been termed class II ERVs. We used PCR amplification and sequencing of retroviral fragments from more than 130 vertebrate taxa to investigate the evolution of the class II retroviruses in detail. We confirm that class II retroviruses are largely confined to mammalian and avian hosts and provide evidence for a major novel group of avian retroviruses, and we identify additional members of both the alpha- and the betaretrovirus genera. Phylogenetic analyses demonstrated that the avian and mammalian viruses form distinct monophyletic groups, implying that interclass transmission has occurred only rarely during the evolution of the class II retroviruses. In contrast to previous reports, the lentiviruses clustered as sister taxa to several endogenous retroviruses derived from rodents and insectivores. This topology was further supported by the shared loss of both the class II PR-Pol frameshift site and the class II retrovirus G-patch domain.     

Isolation of recombinant field strains of Marek's disease virus integrated with reticuloendotheliosis virus genome fragments

Two Marek's disease virus (MDV) field strains were isolated from chickens with tumors independently from Guangdong and Guangxi provinces, and it was confirmed that there were no co-infections with reticuloendotheliosis viruses (REV) in chicken embryo fibroblast cells (CEF) in indirect fluorescence antibody test (IFA) with REV-specific monoclonal antibodies. By dot blot hybridization and PCR of genomic DNA of MDV-infected CEF, it was indicated that LTR fragments of REV genome were integrated into genome of these two MDV field strains. To amplify and clone the integrated REV LTR with MDV sequence at the junction, 4 primers from REV LTR and 7 primers from MDV genome fragment with REV LTR insertion hot points were synthesized and 28 (4x7) pairs of primers (one from REV and another from MDV for each pair) were used in PCR while using the genomic DNA of both strains as the templates. The sequence data demonstrated that both recombinant field strains contained the same REV LTR inserted into MDV at the identical sites in US fragment of the genomes. From the above, it was speculated that both recombinant field MDVs were originated from a same recombinant virus and spread among chicken flocks in two provinces.     

Isolation of recombinant field strains of Marek’s disease virus integrated with reticuloendotheliosis virus genome fragments

Two Marek's disease virus (MDV) field strains were isolated from chickens with tumors independently from Guangdong and Guangxi provinces, and it was confirmed that there were no co-infections with reticuloendotheliosis viruses (REV) in chicken embryo fibroblast cells (CEF) in indirect fluorescence antibody test (IFA) with REV-specific monoclonal antibodies. By dot blot hybridization and PCR of genomic DNA of MDV-infected CEF, it was indicated that LTR fragments of REV genome were integrated into genome of these two MDV field strains. To amplify and clone the integrated REV LTR with MDV sequence at the junction, 4 primers from REV LTR and 7 primers from MDV genome fragment with REV LTR insertion hot points were synthesized and 28 (4x7) pairs of primers (one from REV and another from MDV for each pair) were used in PCR while using the genomic DNA of both strains as the templates. The sequence data demonstrated that both recombinant field strains contained the same REV LTR inserted into MDV at the identical sites in US fragment of the genomes. From the above, it was speculated that both recombinant field MDVs were originated from a same recombinant virus and spread among chicken flocks in two provinces.