Departamento de Bioloxía Fundamental, Universidade de Santiago de Compostela, Spain
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Abstract |
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Introduction |
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In the present work I conducted a comparative sequence approach to reconstruct the evolutionary history of HERV-W, using data from the draft sequence of the human genome. This analysis revealed several unexpected features of HERV-W evolution.
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Materials and Methods |
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HERV-W subfamilies were established by grouping sequences into different sets according to the most variable sites after exclusion of CpG dinucleotide positions (defined using the same criterion as in Costas and Naveira 2000
), with little discrimination for subfamilies because of the fast mutation rate of these dinucleotides to TpG or CpA. Subfamily status was conferred on a sequence set if it was constituted by at least five elements presenting at least two diagnostic nucleotide differences. Subfamily consensus sequences were obtained by choosing the most frequent nucleotide at each position with one exception: those positions considered as CpG in the general alignment were also considered as CpG in the subfamily consensus sequences.
MEGA v2.1 (Kumar et al. 2001
) was used to calculate divergence values within each set of sequences and between different sets of sequences. Net divergence values between different sets of sequences (dN) were calculated by the following formula:
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Results |
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An additional BLAST search was done using as a query the region from the AS3 of HERV-W, located 240 bp upstream of the 3' LTR (Blond et al. 1999
), to the 3' end of the R region, continued by a poly(A) tail. By this strategy, I collected novel 5'-truncated HERV-W retrosequences not recovered in the previous search because of their shorter length. A total of 140 sequences, representing 39 HERV-W proviruses, 40 full-length HERV-W retrosequences, and 61 truncated HERV-W retrosequences, were collected (table 1 ). Furthermore, this search also revealed the existence of solitary R regions with a poly(A) tail flanked by short direct repeats (fig. 1E
), showing that inter-R recombination efficiently removes full-length HERV-W retrosequences from the genome, in a way similar to that giving rise to solitary LTRs from full-length proviruses (Mager and Goodchild 1989
). Besides the known env ORF coding for syncytin (Blond et al. 2000
; Mi et al. 2000
), there are two other HERV-W proviruses preserving ORFs longer than 1,000 bp. One of them, included within the genomic clone NT_022833, extends from amino acids 64 to 524 of syncytin, sharing 87.6% homology with it. The other (NT_006307) is 1,638 bp long, corresponding to the main portion of the pol gene, from the conserved domain 3 of the reverse transcriptase (according to Xiong and Eickbush 1988
) to the end of the RNaseH region.
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Discussion |
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This picture of the intragenomic spread of HERV-W is in clear contrast with other HERV families, such as HERV-K, HERV-H, or ERV9, that remained transpositionally active over extended periods of primate evolution, leading to several distinct subfamilies over time (Anderssen et al. 1997
; Medstrand and Mager 1998
; Costas and Naveira 2000
; Lebedev et al. 2000
). Thus, each HERV family underwent its particular evolutionary history, and these histories may be quite different from each other. The presumably shorter period of amplification in the case of HERV-W (based on the average integration age of the different subfamilies), as well as the apparent lack of intact ORFs, suggests that the MSRV isolated from retroviral particles produced by cell cultures from patients with multiple sclerosis (Perron et al. 1997
) may be an exogenous member of the HERV-W family. The failure to detect intermediate subfamilies between subfamily 1 and subfamily 3 (that present seven diagnostic differences within the U3 region; fig. 2
) also suggests the possibility that these two subfamilies might be originated by two independent germ-line infections.
The most surprising fact of the evolutionary dynamics of HERV-W is the existence of a high proportion of insertions showing characteristic features of retrosequences, such as acquisition of a poly(A) 3' tail, presence of direct flanking repeats of 1016 bp, and a structure resembling mRNAs. Recently, Esnault, Maestre, and Heidmann (2000)
and Wei et al. (2001)
formally disclosed the ability of the non-LTR retrotransposon L1 to retrotranspose polyadenylated RNA transcripts in trans displaying these characteristics. Thus, HERV-W presumably spread by two different mechanisms: (1) the normal retrotransposition process of retroviruses, giving rise to full-length proviruses with intact LTRs, and (2) the parasitism on the L1 element, as in the case of short interspersed elements (SINEs; Mathias et al. 1991
; Ohshima et al. 1996
), giving rise to HERV-W retrosequences. Alternatively, it is legitimate to speculate that the reverse transcriptase of HERV-W itself would be responsible for HERV-W retrosequences formation. Nevertheless, the fact that nonviral RNAs encapsidated in retroviral particles generate integrated cDNA genes lacking the hallmarks of naturally occurring processed pseudogenes (they are 5'- and 3'-truncated and do not contain poly(A) tails) strongly militates against this hypothesis (Dornburg and Temin 1988, 1990
). The existence of both types of elements within each of the subfamilies clearly supports the idea that HERV-W retrosequences formation is dependent on the expression of full-length proviruses, which are the source of genomic RNA. The alternative hypothesis of independent evolution of retrosequences after their origin should give rise to subfamilies constituted only by HERV-W retrosequences, but these subfamilies have not been identified. Taking into account that HERV-W retrosequences are expected to be "dead on arrival" copies, the lower success of HERV-W within the genome, compared with the other afore-mentioned HERV families, might be related to the existence of a considerable proportion of genomic RNA sequestered by the L1 machinery.
The putative impact of HERV-W retrosequences on the genome might be quite different from that of HERV-W proviruses. Retroviral protein expression may cause deleterious effects on the host by several processes. Thus, the antigenic character of proteins encoded by gag and env has been associated with several autoimmune pathologies (Nakagawa and Harrison 1996
; Perron et al. 1997
). The transmembrane domain of the envelope protein presents immunosuppressive effects (Cianciolo et al. 1985
; Haraguchi et al. 1997
), suggesting its possible implication in tumoral processes, leading to the escape of immune rejection by tumoral cells (Mangeney and Heidmann 1998
). Other peptides encoded by small ORFs (two putative small ORFs have been described in HERV-W; Blond et al. 1999
) might interfere with the cellular machinery (Boese et al. 2000
). Furthermore, active proviruses may be the source of new insertions, acting as insertional mutagens (Mitreiter et al. 1994
; Vasicek et al. 1997
). All these deleterious effects are not associated with HERV-W retrosequences, which lack the capability to be expressed because of the loss of LTRs (not only in truncated but also in full-length retrosequences). HERV insertions may also be involved in deleterious chromosomal rearrangements by ectopic recombination between two copies of the same family of HERVs located at different chromosomal loci (Kamp et al. 2000
; Sun et al. 2000
). This effect is expected to be substantially reduced in the case of truncated retrosequences of short length. On the other hand, insertion of HERV-W retrosequences might introduce short enhancer sequences near genes (most of the enhancer signals are within the U3 region), providing raw material for natural selection. Thus, this type of insertion might represent a novel potential mechanism for the evolution of enhancers, adding a new possibility for L1 to shape the mammalian genomes (Kazazian and Moran 1998
; Moran, DeBerardinis, and Kazazian 1999
; Pickeral et al. 2000
).
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Supplementary Material |
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Acknowledgements |
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Footnotes |
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Abbreviations: AS3, splice acceptor site 3; HERV, human endogenous retroviruses; LTR, long-terminal repeat; MSRV, multiple-sclerosis associate retrovirus; NCBI, National Center for Biotechnology Information; ORF, open reading frame.
Keywords: endogenous retrovirus
retrotransposition
HERV-W
L1
MSRV
retrosequence
Address for correspondence and reprints: Departamento de Bioloxía Fundamental, Facultade de Bioloxía, Universidade de Santiago de Compostela, E-15782 Santiago de Compostela, Spain. bfcostas{at}usc.es
.
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