Rex1, together with the related BABAR: elements, represents a new family of non-long-terminal-repeat (non-LTR) retrotransposons from fish, which might be related to the CR1 clade of LINE elements. Rex1/BABAR: retrotransposons encode a reverse transcriptase and an apurinic/apyrimidinic endonuclease, which is very frequently removed by incomplete reverse transcription. Different Rex1 elements show a conserved terminal 3' untranslated region followed by oligonucleotide tandem repeats of variable size and sequence. Phylogenetic analysis revealed that Rex1 retrotransposons were frequently active during fish evolution. They formed multiple ancient lineages, which underwent several independent and recent bursts of retrotransposition and invaded fish genomes with varying success (from <5 to 500 copies per haploid genome). At least three of these ancient Rex1 lineages were detected within the genome of poeciliids. One lineage is absent from some poeciliids but underwent successive rounds of retrotransposition in others, thereby increasing its copy number from <10 to about 200. At least three ancient Rex1 lineages were also detected in the genome project fish Fugu rubripes. Rex1 distribution within one of its major lineages is discontinuous: Rex1 was found in all Acanthopterygii (common ancestor in the main teleost lineage approximately 90 MYA) and in both European and Japanese eels (divergence from the main teleost lineage about 180 MYA) but not in trout, pike, carp, and zebrafish (divergence 100-120 MYA). This might either result from frequent loss or rapid divergence of Rex1 elements specifically in some fish lineages or represent one of the very rare examples of horizontal transfer of non-LTR retrotransposons. This analysis highlights the dynamics and complexity of retrotransposon evolution and the variability of the impact of retrotransposons on vertebrate genomes.

Rex3, the first reverse transcriptase (RT)-encoding retrotransposon isolated from the melanoma fish model Xiphophorus, is a non-long-terminal-repeat element related to the RTE family. The essential features of Rex3 are (1) an endonuclease and a reverse transcriptase, (2) 5' truncations of most of the copies, (3) a 3' tail consisting of tandem repeats of the sequence GATG, and (4) short target site sequence duplications of variable length. Compilation of Rex3 sequences from the pufferfish genome project suggested that, as observed for other members of the RTE family, no additional large open reading frame was present upstream of the endonuclease/reverse transcriptase open reading frame. There are about a thousand copies of Rex3 in the haploid genome of Xiphophorus, some of them probably resulting from recent retrotransposition events. Rex3 RNA was detected by RT-PCR in melanoma and in nontumorous tissues, as well as in melanoma-derived and embryonic cell lines. Rex3 is present in a broad panel of teleost species and was found in the promoter region and in introns of various genes. To our knowledge, Rex3 is the first autonomous retrotransposon described to date which is widespread in teleosts. This wide distribution and occasional association with coding sequences may confer on Rex3 a predisposition to play a role in genome evolution in teleosts.

Abstract:The non-long terminal repeat retrotransposons Rex1 and Rex3 were identified in 13 species of Antarctic fishes from five families of the suborder Notothenioidei. Partial reverse transcriptase gene sequences were characterized for Notothenia coriiceps, Trematomus newnesi and Dissostichus mawsoni (Nototheniidae), and Gymnodraco acuticeps (Bathydraconidae). Rex1 and Rex3 both formed a notothenioidspecific monophyletic group compared to the corresponding elements from other fishes. They globally evolved under purifying selection, showing their activity during notothenioid evolution. Fluorescence in situ hybridization analysis of the chromosomal distribution of Rex1 and Rex3 was performed for several notothenioid fish species. Rex1 was generally less abundant than Rex3, which was widely scattered on the chromosomes with more intense hybridization patterns in some specific zones. Particularly, Rex3 accumulated in Chionodraco hamatus in pericentromeric areas, short arms of some pairs as well as in an intercalary band in the long arm of the Y chromosome similarly to a previously described DNA transposon. Such pattern similarities suggest the presence of autosomal and gonosomal regions of preferential accumulation for different types of repeated elements in notothenioid genomes. To the best of our knowledge, this report is the first description and analysis of retrotransposable elements in Antarctic fish genomes.

All autonomous non-long terminal repeat (non-LTR) retrotransposons reported to date in vertebrates encode an apurinic/apyrimidinic endonuclease-like enzyme necessary for target sequence cleavage and subsequent target-primed reverse transcription. We describe here vertebrate non-LTR retrotransposons encoding another type of endonuclease more related to type IIS restriction enzymes. Such retrotransposons have been detected until now only in trypanosomes, nematodes, and arthropods. The retrotransposon Rex6 was identified in the genome of several teleost fish including Xiphophorus maculatus (platyfish), Oryzias latipes (medakafish), Oreochromis niloticus (Nile tilapia), and Fugu rubripes (Japanese pufferfish). Rex6 encodes a reverse transcriptase and a putative restriction enzyme-like endonuclease and is a member of the R4 family of non-LTR retrotransposons containing the Dong and R4 elements found in nematodes and insects. Rex6 was active in many species during teleost evolution and underwent several bursts of retrotransposition (some of them being relatively recent) leading to a high copy number of Rex6 in the genome of numerous fish. Extremely truncated Rex6-related sequences were detected by database screening in reptiles, including the snake Trimeresus flavoviridis and the lizard Anolis carolinensis, but not in sequences from the human genome project, suggesting that this element might have been lost from certain vertebrate lineages.

The platyfish (Xiphophorus maculatus), in which sex chromosomes are evident from stable and predictable inheritance of sex, is one of the best-studied lower vertebrates with respect to sex determination. In order to identify the structural equivalent for this in the karyotype, which does not contain heteromorphic pairs of chromosomes, two sex-linked molecular probes were used for fluorescent in situ hybridization analysis. One probe, derived from the melanoma oncogene locus ONC-Xmrk, stained both the X and the Y chromosome. This cytogenetic analysis mapped the sex-determining locus to the subtelomeric region of a medium-sized telocentric chromosome. Another probe, a repetitive element (XIR), specifically labeled the Y chromosome in metaphase spreads and in interphase nuclei. The sex chromosomes of X. maculatus can be considered to be at an early stage of evolution of gonosomes. Expansion of the XIR repeat is obviously one of the earliest of the molecular events that lead to divergence of the Y chromosome and recombinational isolation of the sex-determining locus.

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