Identification of a novel family of human endogenous retroviruses and characterization of one family member, HERV-K(C4), located in the complement C4 gene cluster.

We have identified a novel family of about 10-50 human endogenous retrovirus elements (HERVs) and have characterized one family member (HERV-KC4). This retrovirus element is integrated within intron 9 of and complement C4A genes and also in some C4B genes, and is a principal contribution to interlocus and interallelic length heterogeneity of C4 genes. The HERV-K(C4) sequence has a typical retrovirus structure with elements of gag, pol and env domains, flanked by two long terminal repeats (LTRs) and is similar to type A, B and D retroviruses. Multiple termination codons preclude the existence of long open reading frames, suggesting that the HERV-K(C4) sequence is no longer functional. Zoo blot hybridization reveals that New World monkeys appear to lack sequences similar to HERV-K(C4), suggesting that integration has occurred after the divergence of Old and New World monkeys. Retrotransposition of prototype viruses is presumed to have led to the amplification and integration of the members of the family in different loci, which in humans, appear to be dispersed over several chromosomes. The absence of the HERV-K(C4) element in some C4B genes in both humans and orangutangs indicate that the retrovirus inserted into the C4A gene after the duplication of the cluster. Subsequent spread of the HERV-K(C4) sequence to C4B genes presumably occurred by interlocus sequence exchange mechanisms, such as unequal crossover and gene conversion-like mechanisms.     

Nucleotide sequence of the Syrian hamster intracisternal A-particle gene: close evolutionary relationship of type A particle gene to types B and D oncovirus genes.

We determined the complete nucleotide sequence of the intracisternal A-particle gene, IAP-H18, cloned from the normal Syrian hamster liver DNA. IAP-H18 was 7,951 base pairs in length with two identical long terminal repeats of 376 base pairs at both ends. On the coding strand, imperfect open reading frames corresponding to gag and pol of the retrovirus genome were observed, whereas many stop codons were present in the region corresponding to env. The putative H18 gag gene (809 amino acids) had a sequence homologous to the N-terminal half of the mouse mammary tumor virus gag gene and locally to the Rous sarcoma virus gag gene. The putative H18 pol gene (900 residues) was homologous to the Rous sarcoma virus pol gene almost throughout the entire region. Two conserved regions among the retrovirus pol genes have been reported. One presumably corresponds to the DNA polymerase and the RNase H domain, and the other corresponds to the DNA endonuclease domain of the multifunctional protein pol. By the comparison of the deduced amino acid sequences of the putative endonuclease domain of six representative oncovirus genomes, a phylogenetic tree of the oncovirus genomes was constructed, and the intracisternal A-particle (type A) genome was found to be more closely related to the mouse mammary tumor virus (type B) and squirrel monkey retrovirus (type D) genomes.     

Novel Endogenous Type C Retrovirus in Baboons: Complete Sequence, Providing Evidence for Baboon Endogenous Virus gag-pol Ancestry

A complete endogenous type C viral genome has been isolated from a baboon genomic library. The provirus, Papio cynocephalus endogenous retrovirus (PcEV), is 8,572 nucleotides long, and 38 to 59 proviral copies per baboon genome are found. The PcEV provirus possesses the typical simple retroviral gene organization, including two long terminal repeats and genes encoding gag, pol, and env proteins. The open reading frames for gag-pol and env are complete but have premature stop codons or frameshift mutations. The primer binding site of PcEV is complementary to tRNAGly. The gag and pol genes of PcEV are closely related to those of the baboon endogenous virus (BaEV). The env coding region of PcEV is related to the env genes of type C retroviruses. This suggests that PcEV is one of the ancestors of BaEV contributing the type C gag-pol genome fragment to the type C/D recombinant virus BaEV. Earlier it was shown that another endogenous type D virus (simian endogenous retrovirus) provided the env gene for BaEV (A. C. van der Kuyl et al., J. Virol. 71:3666-3676, 1997).     

Nucleotide sequence of a complete mouse intracisternal A-particle genome: relationship to known aspects of particle assembly and function.

The 7,095-nucleotide sequence of a mouse genomic intracisternal A-particle (IAP) element, MIA14, is reported. MIA14 is known to be colinear with IAP 35S RNA and to contain functional long terminal repeats. Its internal genetic organization was determined by comparisons with a homologous Syrian hamster element and the related retroviruses simian retrovirus 1 (simian type D) and Rous sarcoma virus (avian type C). MIA14 contains a gag-protease open reading frame of 827 codons and a pol region of 867 codons entered by a frame shift of -1. The env region of 1,100 base pairs has multiple stop codons in all reading frames, consistent with the failure thus far to detect IAP-related glycosylated envelope components. RNA transcribed in vitro from a cDNA clone containing a closely homologous gag-protease open reading frame was translated in a cell-free system. The main product was a 73-kilodalton polypeptide immunoprecipitable with antiserum against the authentic IAP gag-related structural protein p73. Rather than ending at the gag-protease boundary, p73 appears to contain 7 to 8 kilodaltons of peptide encoded by the protease domain, a peculiarity possibly related to the observed impairment of normal protein processing in IAPs. The N-terminal 217 codons of gag are unique to murine IAPs and may have been contributed by recombination with a cellular gene. The mouse-specific region of gag encodes a hydrophobic signal peptide with an atypical cleavage site. Delayed cleavage of this peptide could result in anchoring of newly synthesized p73 to the endoplasmic reticulum membrane and restriction of particle assembly to this site.     

Characterization of the bovine endogenous retrovirus beta3 genome

We recently used degenerate PCR and locus-specific PCR methods to identify the endogenous retroviruses (ERV) in the bovine genome. Using the ovine ERV classification system, the bovine ERVs (BERVs) could be classified into four families. Here, we searched the most recently released bovine genome database with the partial nucleotide sequence of the pro/pol region of the BERV beta3 family. This allowed us to obtain and analyze the complete genome of BERV beta3. The BERV beta3 genome is 7666 nucleotides long and has the typical retroviral organization, namely, 5'-long terminal repeat (LTR)-gag-pro-pol-env-LTR-3'. The deduced open reading frames for gag, pro, pol and env of BERV Beta en- code 507, 271, 879 and 603 amino acids, respectively. BERV beta3 showed little amino acid similarity to other betaretroviruses. Phylogenetic analysis showed that it clusters with HERV-K. This is the first report describing the genetic structure and sequence of an entire BERV.     

Inversion of a chicken ets-1 proto-oncogene segment in avian leukemia virus E26.

The segment of the avian leukemia virus E26 genome near the termination of the p135gag-myb-ets open reading frame contains an inversion of the chicken ets-1 sequence. The inversion contains at least 41 bp and may be as large as 46 bp. This results in the replacement of 13 amino acids of chicken ets-1, with 16 amino acids derived from reverse complement of the normal ets-1 coding strand or read-through into E26 env sequences. At least 13 of these codons are specified by the inverted ets sequences. This represents the first reported occurrence of inverted oncogene sequences in a natural retrovirus. The inverted ets sequences are immediately followed by sequences homologous to the Rous sarcoma virus Prague B env gene. Since the E26 env sequence is more closely related to subgroup B avian retroviruses than to avian retroviruses from subgroups A, C, D, or E, the progenitor of E26 was a virus belonging to avian retrovirus subgroup B.     

The human endogenous retrovirus family HERV-K(HML-3)

A substantial amount of the human genome is composed of human endogenous retroviruses (HERVs). Manifold HERV families have been identified, among them several so-called HERV-K(HML) families. Although the HERV-K(HML-2) family has been studied in detail, other HERV-K families are not as well characterized. We describe here the HERV-K HML-3 family in more detail. We estimate that there are about 140 proviral loci or remains of such per haploid genome. Most loci are severely mutated. Proviruses displaying larger deletions in gag and pol are common. A multiple alignment of 73 HERV-K(HML-3) sequences displays several potentially important differences compared with the HERVK9I sequence in Repbase. A consensus sequence with open reading frames for all retroviral genes was generated, for which intact dUTPase motifs and env gene variants with different coding capacities are observed. Phylogenetic analysis shows near-monophyly with distinction of two closely related subgroups. Proviruses formed about 36 million years ago. However, no continuous activity through primate evolution is indicated.     

Complete nucleotide sequence and genome organization of a Drosophila transposable genetic element, 297

The complete nucleotide sequence of 297, a Drosophila copia-like transposable element, was determined and compared with those of other similar Drosophila elements and mammalian retrovirus proviruses. It was found that 297 contains three long open reading frames, comparable in sizes and locations with gag, pol, and env genes in the proviruses of replication-competent retroviruses in vertebrates. The first and second open reading frames of 297 exhibit sequence homologies to gag and pol, respectively, of Moloney murine leukaemia virus. In particular, as with 17.6, another Drosophila copia-like element, the second open reading frame of 297 was shown to be very similar in its entire organization to the retroviral pol gene and to consist of three enzymatic domains. By contrast, no appreciable homology was found between the third open reading frame of 297 and the retroviral env gene. It is also suggested that 297 and 17.6 are a peculiar pair of copia-like elements recently diverged from a common progenitor.     

Nucleotide sequence of the intracisternal A-particle genome inserted 5'' to the interleukin-3 gene of the leukemia cell line WEHI-3B.

The nucleotide sequence of the intracisternal A-particle genome IAP-IL3 is presented. This IAP element was found to have inserted upstream of the promoter of the interleukin-3 gene of the leukemia cell line WEHI-3B. IAP-IL3 is 5095 bp in length, with identical long terminal repeats (LTRs) of 337 bp. The LTRs show many of the conserved sequence elements identified in other retroviruses. Comparison with other available sequences of IAP genomes indicates that IAP-IL3 is a deleted type I element. It carries a deletion covering the 3' end of the putative IAP gag gene and extending into the 5' end of the putative IAP pol gene. IAP-IL3 has extensive sequence homology with an IgE-binding factor cDNA and evidence is presented indicating that it was derived from a member of the mouse IAP sequence family. Comparison between the pol region of IAP-IL3 and other retroviruses suggests that IAP-IL3 is most closely related to type B and type D retroviruses.     

Human endogenous retrovirus family HERV-K(HML-5): status, evolution, and reconstruction of an ancient betaretrovirus in the human genome

The human genome harbors numerous distinct families of so-called human endogenous retroviruses (HERV) which are remnants of exogenous retroviruses that entered the germ line millions of years ago. We describe here the hitherto little-characterized betaretrovirus HERV-K(HML-5) family (named HERVK22 in Repbase) in greater detail. Out of 139 proviruses, only a few loci represent full-length proviruses, and many lack gag protease and/or env gene regions. We generated a consensus sequence from multiple alignment of 62 HML-5 loci that displays open reading frames for the four major retroviral proteins. Four HML-5 long terminal repeat (LTR) subfamilies were identified that are associated with monophyletic proviral bodies, implying different evolution of HML-5 LTRs and genes. Sequence analysis indicated that the proviruses formed approximately 55 million years ago. Accordingly, HML-5 proviral sequences were detected in Old World and New World primates but not in prosimians. No recent activity is associated with this HERV family. We also conclude that the HML-5 consensus sequence primer binding site is identical to methionine tRNA. Therefore, the family should be designated HERV-M. Our study provides important insights into the structure and evolution of the oldest betaretrovirus in the primate genome known to date.     

Discovery of a New Endogenous Type C Retrovirus (FcEV) in Cats: Evidence for RD-114 Being an FcEVGag-Pol/Baboon Endogenous Virus BaEVEnv Recombinant

Analysis of a cat genomic DNA library showed that cats harbor a previously unrecognized endogenous type C retrovirus, whose env gene has homology to the murine Fv-4 resistance gene. This unique retrovirus, designated FcEV (Felis catus endogenous retrovirus), has a type C pol gene, closely related to the primate Papio cynocephalus endogenous virus (PcEV) pol, not overlapping the env gene, unlike in other type C retroviruses, and is presumably present in a higher copy number than RD-114. Phylogenetic analysis of FcEV and RD-114 fragments amplified from cat species and comparison with baboon endogenous virus (BaEV) fragments from monkeys suggested that RD-114 does not represent the cat strain of BaEV but is actually a new recombinant between FcEV type C genes and the env gene of BaEV. Although BaEV did appear to have infected an ancestor of the domestic cat lineage, it was a de novo recombinant that made its way into the cat germ line.     

Proviral genome of radiation leukemia virus: molecular cloning of biologically active proviral DNA and nucleotide sequence of its long terminal repeat.

The proviral genome of a leukemogenic and thymotropic C57BL/Ka mouse retrovirus, RadLV/VL3(T+L+), was cloned as a biologically active PstI insert in the bacterial plasmid pBR322. Its restriction map was compared with those, already known, of two nonthymotropic and nonleukemogenic viruses of the same mouse strain: the ecotropic BL/Ka(B) virus and the xenotropic constituent of the radiation leukemia virus complex. Differences were observed around the gag-pol gene junction, in the pol gene, and in the env gene. Moreover, the nucleotide sequence of the RadLV/VL3(T+L+) long terminal repeat revealed the existence of two copies of a 43-base-pair sequence, of which BL/Ka(B) possesses only one copy.     

Complete nucleotide sequence of simian endogenous type D retrovirus with intact genome organization: evidence for ancestry to simian retrovirus and baboon endogenous virus.

A complete endogenous type D viral genome has been isolated from a baboon genomic library. The provirus, simian endogenous retrovirus (SERV), is 8,393 nucleotides long and contains two long terminal repeats and complete genes for gag, pro, pol, and env. The primer binding site is complementary to tRNA(Lys)3, like in lentiviruses. The env GP70 protein is highly homologous to that of baboon endogenous virus (BaEV). PCR analysis of primate DNA showed that related proviral sequences are present in Old World monkeys of the subfamily Cercopithecinae but not in apes and humans. Analysis of virus and host sequences indicated that the proviral genomes were inherited from a common ancestor. Comparison of the evolution of BaEV, exogenous simian retrovirus types 1 to 3 (SRV1 to SRV3), and SERV suggests that SERV is ancestral to both BaEV and the SRVs.