* Laboratories of Genome Dynamics, Center of Anatomy and Cell Biology, Medical University of Vienna, Vienna, Austria; and Children's Cancer Research Institute, Vienna, Austria
Correspondence: E-mail: sylvia.hagemann{at}meduniwien.ac.at.
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key Words: Drosophila P transposons Homo sapiens Danio rerio Gallus gallus THAP proteins molecular domestication
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In humans, at least 45% of the genome belong to transposable elements, and a number of single-copy genes seem to have originated from them. Until now, 48 domesticated human genes probably originating from up to 39 different transposon copies could be identified (Hagemann and Pinsker 2001; International Human Genome Sequencing Consortium 2001; Nekrutenko and Li 2001). Most of them originated from DNA transposons, although only about 6% of the human transposable elements belong to this transposon type.
In 2001 we reported the detection of a P-homologous sequence in the human genome (Hagemann and Pinsker 2001), which we subsequently named Phsa (P homolog of Homo sapiens). From the sequences available at that time, we concluded that Phsa codes for a protein of 759 amino acids. Closer examination of the genomic organization of Phsa as well as the comparison with several insect P element sequences revealed the absence of transposon characteristic terminal inverted repeats and led to the assumption that Phsa is a stationary sequence in the human genome (Hagemann and Pinsker 2001). Through database searches, P-homologous sequences have been identified also in the genomes of chicken (Gallus gallus) and cattle (Bos taurus) (Hagemann and Pinsker 2001), indicating a widespread occurrence in vertebrates.
Roussigne et al. (2003a) described a novel proapoptotic factor, which they designated THAP1. Database searches with both the nucleotide and the amino acid sequences of human THAP1 revealed that the first 89 N-terminal amino acid (aa) residues constitute a novel protein motif, the THAP domain, which seems to be evolutionary conserved and restricted to animals. They suggested that THAP domain proteins might belong to a new family of cellular DNA-binding proteins (Roussigne et al. 2003b) and demonstrated that the THAP domain shows striking similarities to the site-specific DNA-binding domain of the D. melanogaster P element transposase. They designated this P element transposasehomologous protein THAP9 (THAP domain containing 9), which corresponds to the putative Phsa gene product detected in 2001 (Hagemann and Pinsker 2001) and further characterized in this paper.
In the present study, we prove experimentally that the human Phsa is a single-copy gene located on the long arm of chromosome 4. Pgga (P homolog of Gallus gallus), the homologous gene from chicken, as well as a mouse and a rat P-homologous rudiment, can be found at orthologous positions. With respect to zebrafish, Blast searches resulted in the detection of a considerable number of P-homologous sequences (Pdre [P homolog of Danio rerio]) with DNA-transposon characteristic structural features such as terminal inverted repeats and target-site duplications, strongly indicating that they were mobile within the zebrafish genome. To gain some insights into the possible function of the domesticated Phsa, we carried out expression studies to characterize the transcription pattern in different human tissues. Finally, we describe the phylogenetic relationship of Phsa-homologous sequences of various vertebrates and discuss their evolutionary history as active transposable elements versus domesticated genes.
![]() |
Materials and Methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
|
Fluorescence in situ Hybridization (FISH) Analysis
To identify the location of the Phsa gene and its possible homologs on human chromosomes, FISH using a locus-specific probe was performed. For this purpose, a 15-kb lambda phage clone covering the 3' end of Phsa that was obtained by screening a human genomic library was used. The probe was labeled with DIG-II-dUTP by nick translation, and FISH was carried out as previously described in Koenig et al. (2002). For immunodetection of the probe, sheep antidigoxigenin FITC (1:100) (Roche Diagnostics, Mannheim, Germany) and rabbit antisheep FITC (1:100) (Dako, Hamburg, Germany) were used.
Genome-Walking Analysis in Chicken
To determine the copy number of P-homologous sequences within the genome of chicken, genome-walking experiments were carried out using the Universal Genome Walker Kit of CLONTECH Laboratories (BD Biosciences, San Jose, Calif.). Genomic DNA was digested with the restriction enzymes DraI, EcoRV, PvuII, and StuI and genome walking was performed towards the 3' end of the target gene following the manufacturer's instructions. The nested primers Pgga+3 and Pgga+4 served as gene-specific primers, binding in exon 2 of the Pgga gene (table 1 and fig. 1B). The obtained PCR products were cloned using the pGEM-T Easy vector system and subjected to automated sequencing.
Quantitative Real-Time PCR
To examine the levels of specific transcripts of Phsa in different human tissues, multiple tissue cDNA panels (Human MTC Panel [BD Biosciences, San Jose, Calif.]) were analyzed. These cDNA panels contain normalized, first-strand cDNA preparations from RNA of the following pooled human tissues or cell lines that were derived from at least two male and/or female Caucasians: brain (whole), heart, kidney, liver, lung, pancreas, peripheral blood leukocyte, placenta, and skeletal muscle (for details regarding the normalization procedure, see the cDNA Panels User Manual). The human cDNA samples were subjected to quantitative real-time PCR (qPCR). A 303-bp fragment of exon 5 of Phsa was amplified with the primers q1+Phsa and q1Phsa (table 1) using the LightCycler DNA Master SYBR Green I kit (Roche Diagnostics, Mannheim, Germany). In this assay, the housekeeping gene (HK) for glycerol-3-phosphate dehydrogenase (G3PDH, EC 1.1.1.8) served as positive control, and the respective qPCR was conducted with the primer pair q1+G3PDH and q1G3PDH (table 1), which lead to a 318-bp PCR product. The qPCR was performed on the LightCycler instrument (Roche Diagnostics, Mannheim, Germany) in 15-µl reactions containing 1.5 µl FastStart DNA Master reaction mix, 0.65 µM MgCl2, 5 pM of each primer, and 1 ng/µl normalized cDNA sample. The PCR conditions were 95°C for 10 min, followed by 45 cycles at 95°C for 10 s, 59°C (Phsa and G3PDH) for 5 s, and 72°C for 20 s (slopes were 20°C/s). Fluorescence was measured at the end of the extension phase. To confirm the specificity of the amplified products, melting curves were performed at the end of the amplification by cooling the samples at 20°C/s to the respective annealing temperatures of both primer pairs and then increasing the temperature to 95°C at 0.2°C/s with fluorescence measurement every 0.1°C. Two standard curves for Phsa and G3PDH, respectively, were generated using serial dilutions of the liver sample of the cDNA panel at concentrations of 0.008, 0.01, 0.013, 0.02, 0.05, 0.1, 0.2, and 1 ng/µl. All standards were amplified in duplicates and a regression curve was computed, which served as internal reference for further calculations. The identical cDNA samples were examined at four different times, and in each experiment, the samples were measured in duplicates. For the statistical analysis, means, standard deviations, and standard errors were computed. The relative abundance of Phsa gene transcript was measured by calculating the ratio of mean concentrations of this gene over the mean concentrations of G3PDH for the respective tissue samples. Differences between Phsa and G3PDH synthesis were evaluated by one-way analysis of variance (1-way ANOVA), followed by the paired samples (two sample) t-test, respectively, both as implemented in Analyse-it version 1.71 (Analyse-it Software Ltd.) for Microsoft Excel.
Phylogenetic Analysis of P-Homologous Sequences
Details regarding the 26 vertebrate species analyzed in this study are given in table 2. Total genomic DNA of the eight primates was provided by the German Primate Center (Göttingen, Germany), commercially available DNA was used for an additional eight vertebrates (EMD Biosciences, San Diego, Calif.), and the DNA of the remaining nine specimens was extracted following the protocol described in the Genetic Analysis Manual (LI-COR, Inc. 1999). Depending on the quality and amount of genomic DNA, 1 to 5 µl (1 µg) were subjected to PCR amplification using primers Pver1+ and Pver1 (table 1). PCR reactions were electrophoresed on 1% agarose/EtBr gels and blotted onto nylon membranes (Amersham Biosciences, Uppsala, Sweden) by Southern transfer following standard protocols (Sambrook, Fritsch, and Maniatis 1989). For hybridization, a 700-bp fragment of exon 5 of Phsa was amplified with the primers Pver1+ and Pver1 and cloned using the pGEM-T Easy vector system (Promega, Madison, Wis.). This probe will be referred to as Phsa700-C (fig. 1A). After PCR DIG labeling, this probe was used to identify homologs of Phsa with the DIG detection system. Positive PCR fragments were cloned using the pGEM-T Easy vector system and subjected to automated sequencing. P-homologous DNA sequences (719 bp) and the deduced amino acid sequences (239 aa) of 21 vertebrate species (table 2) were aligned by the program ClustalX (Thompson et al. 1997). A maximum-likelihood (ML) analysis was conducted by application of the quartet-puzzling approach of Tree-Puzzle (Schmidt et al. 2002). The substitution process was modeled by implementing the HKY85 model (Hasegawa, Kishino, and Yano 1985) for the DNA sequence data, and the Dayhoff model (Dayhoff, Schwartz, and Orcutt 1978) for the protein data, respectively. The DNADIST option of PHYLIP (Felsenstein 2004) was used to perform a Neighbor-Joining analysis (NJ [Saitou and Nei 1987]) based on the F84 model (Felsenstein and Churchill 1996) of nucleotide evolution and the JTT92 model (Jones, Taylor, and Thornton 1992) of amino acid sequence evolution. To evaluate the quality of the phylogenetic signal of Phsa for resolving phylogenetic relationships among vertebrates, the obtained Phsa gene trees were compared with those of the mitochondrial (mt) cytochrome b (cyt b). Therefore, the entire mtDNA cyt b gene DNA sequences (1,149 bp) and the deduced amino acid sequences (382 aa) of 25 vertebrate species (table 2) were aligned with the program ClustalX (for accession numbers, see Supplementary Material). The mtREV24 model (Adachi and Hasegawa 1996) and the JTT92 model of amino acid sequence evolution, as well as the TrN model (Tamura and Nei 1993) and the F84 model of nucleotide evolution, were used for likelihood and distance analysis. The robustness of the phylogenies was assessed by the bootstrap percentage, and by the reliability percentages, (i.e., the number of times the group appears after 10,000 puzzling steps).
|
![]() |
Results and Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
Genomic Organization of the P-Homologous Pgga in Gallus gallus
In 2001, we reported the discovery of a P-homologous sequence in chicken (Hagemann and Pinsker 2001), which will be referred to as Pgga. The genome-walking experiments revealed a single copy of Pgga, which corresponds to the sequence represented by accession number NW_060347, confirming the Blast search results suggesting that Pgga is a single-copy gene like its human homolog, Phsa. The P-homologous region extends from nt 16680605 to nt 16676896, comprises a region of 3,710 bp, and is located on the long arm of chromosome 4. Based on the alignment of the cDNA sequence (accession number XM_420555) with the corresponding genomic sequence, we were able to determine the exon/intron limits of this gene. Pgga consists of one intron (ig = 1,185 bp) and two exons (eg1 = 156 bp, eg2 = 2,369 bp [fig. 1B]), which can be translated into a P elementhomologous protein of 681 amino acid residues. The intron belongs to the minor group of GCAG introns, the 5' splice donors of which are thought to play an important factor in the regulation of alternative splicing (Farrer et al. 2002). Exon eg1 is homologous to the human exon eh4, whereas eg2 is homologous to the human eh5 (fig. 1A and B), and the start codon of Pgga corresponds to the previously assumed start codon of the first human cDNA entry (accession number AK026973).
In comparison with the putative Phsa protein, the presumed Pgga gene product shows a N-terminal truncation, thus, lacking the DNA-binding THAP domain (fig. 1A and B and fig. 4A and B). Downstream of Pgga, the functional gene LOC422595 (cDNA accession number XM_420554) was identified, giving rise to a protein similar to the hypothetical protein DKFZp686L1814 located downstream of Phsa in human (fig. 1D). These findings strongly support the orthologous positions of Phsa and Pgga in their respective host genomes (fig. 1C and D).
Genomic Organization of P-Homologous Pdre Elements in Danio rerio
Blast searches with the deduced amino acid sequence of Phsa as query sequence revealed four different Blast hits in the zebrafish genome. The homologous sequences are located in linkage group 2 (accession number BX511023), group 4 (accession number BX890548), group 17 (accession number BX324003), and group 22 (accession number CR388079) and will be referred to as Pdre2, Pdre4, Pdre17, and Pdre22. Multiple alignment analyses of these sequences showed that the repeated region of Pdre2 in linkage group 2 is the most extended one. Consequently, this sequence was used for further Blast searches, leading to the detection of at least 50 P-homologous sequences spread throughout the genome of zebrafish. Most of them are internally deleted copies (e.g., in linkage groups 1 [accession number BX537346: nt 47346 to nt 48360], 3 [accession number AL929078: nt 95952 to nt 96777], and 20 [accession number AL954815: nt 148549 to nt 149356]) referred to as Pdre1, Pdre3, and Pdre20, whereas Pdre4 (accession number BX890548: nt 39502 to nt 47547), Pdre17 (accession number BX324003: nt 50600 to nt 56639), and Pdre22 (accession number CR388079: nt 17390 to nt 27566) are terminally truncated. One putative complete transposon was found in linkage group 2 (Pdre2 [fig. 3]), which is 11,149 bp long (accession number BX511023: nt 198423 to nt 209571), possesses 13-bp terminal inverted repeats (5'-CATACCTGTCAAC-3'/5'-GTTGACAGGTATG-3') together with 12-bp subterminal inverted repeats (5'-TGTTTAAACCAA-3'/5'-TTGGTTTAAACA-3'), and is flanked by an 8-bp target-site duplication (5'-AGGTGAAT-3'). Terminal inverted repeats as well as the Drosophila P transposontypical 8-bp target site duplications can also be found for the analyzed internally deleted elements Pdre1 (5'-CCTTTAAG-3'), Pdre3 (5'-CATCCAAC-3'), and Pdre20 (5'-GTCTACAT-3'), but they are missing for the terminally truncated elements Pdre4, Pdre17, and Pdre22. The large sizes of Pdre2 (11,149 bp) and of the terminally truncated elements Pdre4 (8,046 bp), Pdre17 (5,940 bp), and Pdre22 (10,177 bp) indicate that this is a conserved feature of Pdre elements.
|
With respect to Pdre2, the only long open reading frame (ORF) extends from nt 3448 to nt 6078 comprising a region of 2,631 bp that can be translated into a P elementhomologous protein of 877 amino acid residues. Because a 5' start codon is lacking within this ORF, we performed translation product analyses of the upstream region by searching for conserved protein domains (Marchler-Bauer et al. 2003). Using the deduced amino acid sequence from the first reading frame as query sequence revealed a THAP domain (E-value: 2 x1004), the putative DNA-binding domain as described by Roussigne et al. (2003b). The THAP domaincorresponding DNA sequence comprises the Pdre2 region from nt 1639 to nt 1806, but the N-terminal region of the THAP domain was missing. Therefore, the upstream region was searched for additional exons and a further small exon could be detected comprising the Pdre2 sequence from nt 849 to nt 880, suggesting a transposase gene consisting of three exons (eda1 = nt 849 to nt 880, eda2 = nt 1656 to nt 1808, and eda3 = nt 3468 to 6078) and two introns (ida1 and ida2 [fig. 3]). Both introns belong to the type of GCAG introns indicating that the Pdre2 gene could be alternatively spliced (Farrer et al. 2002). Based on these findings, we constructed a hypothetical mRNA, which could be translated into a protein of 932 amino acid residues with a N-terminal THAP domain (E-value: 7x1008) overlapping the eda1/eda2/eda3 boundaries; hence, it follows that our splicing interpretation is correct.
In contrast to Pdre2, the ORFs of the putative exon eda3 of the terminally truncated elements Pdre4, Pdre17, and Pdre22 are destroyed. Thus, within the zebrafish genome, two scenarios can be observed for the P-homologous sequences: Pdre4, Pdre17, and Pdre22 are terminally truncated and, therefore, present immobile components of the zebrafish genome, whereas Pdre2 and the analyzed internally deleted copies Pdre1, Pdre3, and Pdre20 have terminal inverted repeats and are flanked by target-site duplications, both typical characteristics for mobile DNA transposons.
Protein Domains of Vertebrate P-Homologous Sequences
Roussigne et al. (2003a) described a novel protein motif, the THAP domain, which shows striking similarities to the DNA-binding site of Drosophila P element transposases. To date, 113 THAP domaincontaining proteins are listed in the databases, including seven murine and 12 human THAP proteins, and two of them were known to have metabolic functions (Gale et al. 1998; Smit 1999; Roussigne et al. 2003a). The THAP motif contains a putative DNA-binding domain that is characterized by tetrahedral formation of one or two zinc ions by conserved cysteine and histidine residues. Zinc-coordinating proteins constitute the largest group of transcription factors in eukaryotic genomes and were also found in domains that mediate proteinprotein interactions. Except for the D. melanogaster protein CG10631 (accession number AAF53840), in which 27 THAP domains occur consecutively, nearly all known THAP domains are located in the N-terminus of the protein, covering about 90 amino acid residues. In figure 4A, the THAP-domain consensus sequence is compared with the corresponding regions of the P-homologous proteins of the fruit fly, human, and zebrafish. The conserved sequence motif Cys-Xaa24-Cys-Xaa3550-Cys-Xaa2-His, together with the three strictly conserved residues P, W, and F (consensus positions 23, 32, and 55 in figure 4A) (Roussigne et al. 2003b), can be found in all three proteins. The C-terminal AVPTIF box is highest conserved in the putative Pdre2 protein, whereas several amino acid substitutions occur in the Drosophila P-element transposase and in the Phsa protein (fig. 4A).
The alignment of the P-homologous proteins from human, chicken, and zebrafish as presented in figure 4B shows the absence of the THAP domain in the putative Pgga protein. As the presumed start codon of Pgga corresponds to the previously postulated start codon of the first Phsa-cDNA entry (accession number AK026973), it can be speculated that the respective cDNAs are alternative splicing products and that the THAP domaincontaining Pgga transcript could not be detected so far. In the case that the THAP domain is really deleted in the Pgga protein, the DNA-binding domain would have been lost, indicating an altered function of the Pgga gene product within chicken.
Another known P-transposase motif, the leucine zipper, is supposed to play a transposition inhibitory function in Drosophila because of dimerization between the transposase and its repressor (Lee, Mul, and Rio 1996). This motif can be found in all Drosophila P elementrelated proteins and comprises the region from aa 101 to aa 122 of the D. melanogaster transposase (accession number A24786 [Rio 1990]). This leucine zipper is lacking not only in the domesticated P-homologous proteins of human and chicken but also in the presumable transposable zebrafish elements. On the assumption that leucine zipper domains are mediating the repression of transposition, the lack of this domain indicates that activation and repression within the zebrafish genome might follow another mechanism. DNA methylation and demethylation, as well as RNA interference, possibly mediate transposon silencing in various eukaryotic genomes (Kato et al. 2003; Zhou, Cambareri, and Kinsey 2001; Montgomery 2004; Novina and Sharp 2004) and could be the silencing mechanism of Pdre elements in the zebrafish genome, too.
Expression Analysis of Phsa
To characterize the tissue-specific gene expression pattern of Phsa, cDNA panels that comprise cDNA pools of various tissues, including brain (whole), heart, kidney, liver, lung, pancreas, peripheral blood leukocyte, placenta, and skeletal muscle, were subjected to quantitative real-time PCR. The mean calculated concentration of the Phsa transcript in the various tissues ranged from 7.983x108, as observed in brain, to 1.542x1012, as found in the peripheral blood leukocyte sample (fig. 5A). For the HK gene, G3PDH the highest level of gene activity was detected in skeletal muscle (3.635x1015), whereas it was the lowest in placenta (1.8x1011 [fig. 5A]). The significance of the high level of G3PDH expression in skeletal muscle was confirmed by 1-way ANOVA (2-tailed, P = 0.0012 to 0.0013; t statistic 3.27 to 3.22 [data not shown]) as might be expected for an enzyme like G3PDH, possible because of the high ATP production in this tissue.
|
|
Molecular Phylogeny of P-Homologous Vertebrate Sequences
By means of PCR amplification or Blast searches, we were able to characterize P-homologous DNA sequences of almost all analyzed vertebrates (table 2). By using the primer pair Pver1+ and Pver1, we detected no P-homologous DNA sequences in the five rodent species examined: house mouse (Mus musculus), black rat (Rattus rattus), brown rat (Rattus norvegicus), root vole (Microtus oeconomus), and the European beaver (Castor fiber). Based on paleontological records, a divergence time of about 310 Myr for the separation of birds and mammalian lineages can be assumed, whereas the radiation of the recent mammalian orders dates back approximately 100 Myr (Nelson 1996; Nei and Glazko 2002). Hence, the rodent sequence should have diverged to a lesser extent from other mammals than did the latter from chicken. However, studies focusing on the molecular phylogeny of vertebrates, emphasizing the mammals, argued for an accelerated evolution of rodent genes compared with primate genes (Nei and Glazko 2002; Cotton and Page 2002). To account for the possibly more divergent P-homologous sequences in rodents, we used further primer combinations as well as touchdown PCR. Because of the fact that we were able to isolate and detect the highly divergent P element homolog from the chicken genome and in zebrafish, respectively, but not from any rodents studied so far, we supposed that the functional P-homologous sequence has been lost in the rodent lineage. This interpretation was confirmed by using the UCSC genome browser (mouse assembly, May 2004). On the mouse chromosome 5, positive Blast hits comprised the region from nt 22932957 to nt 22933964 (accession number NT_039308). This sequence corresponds partially to the 3'untranslated region of the Phsa gene, suggesting that it is a P-homologous rudiment in mouse. Using the cDNA (accession number BX640657), which was used to confirm the orthologous positions of Phsa and Pgga, as query sequence Blast search revealed that the P-homologous rudimentary sequence is located at orthologous position in the mouse genome, too (accession number NT_039308: nt 22939616 to nt 22996577). The same sequence arrangement can be found in rat, where the P-homologous rudiment (accession number AABR03089973: nt 27116 to 26115) and the neighboring gene (accession number AABR03089973: nt 1 to nt 17906) are located on chromosome 14. In conclusion, our results suggest that the failure to amplify P-homologous sequences in rodents is more likely caused by gene loss than by sequence divergence in this group.
Based on the obtained DNA sequences and their deduced amino acid alignment, we reconstructed the phylogenetic history of the analyzed vertebrate P-homologous elements and compared the results with those of the phylogenetic relationships among vertebrates inferred from mt cyt b sequences (table 4 and fig. 6A and B). The reconstructed topologies for the deduced P-homologous and the cyt b amino acid data sets are shown in figure 6. The reconstructed phylogenies of both approaches revealed congruence for each data set, although only the trees of the ML analyses of the amino acid data set of each gene are presented (fig. 6A and B). Comparing the ML tree derived from P homologs with that from cyt b, the topologies were not congruent. Moreover, the tree search for the cyt b protein data set revealed a better resolution of the computed tree (i.e., lower number of unresolved quartets [table 4]). Both gene trees clearly confirmed the monophyly of the primates and supported the paraphyletic situation of the insectivores, as well as the aberrant position of the ring-tailed lemur, Lemur catta. In the P-homologous ML tree (fig. 6A), the carnivores formed a monophyletic group, which could not be confirmed by the cyt b ML analysis (fig. 6B). Interestingly, the cyt b ML tree displayed a paraphyletic positioning of the rodents (fig. 6B). With respect to these results, we can conclude that our P-homologous sequence data reflect to a certain extent the phylogenetic relationships among vertebrates (i.e., ML analyses of entire mitochondrial genomes [Arnason et al. 2002; Nikaido et al. 2001]). In conclusion, the phylogeny of the P-homologous sequences from the vertebrates studied so far is more or less in accordance with the species phylogeny, indicating their vertical transmission.
|
|
In zebrafish, P-homologous sequences are spread throughout the genome, indicating their transposable activity and contributing to two different scenarios that can be observed within the zebrafish genome: the terminally truncated Pdre sequences represent immobile components of the genome, whereas Pdre2 and the analyzed internally deleted copies show structural features typical for DNA transposons. Quite contrary features can be described for Phsa in human and Pgga in chicken, where they represent stationary single-copy genes. Phsa, Pgga, and the P-homologous mouse and rat rudiments are located at orthologous positions, thus, favoring the "source gene hypothesis." Although the immobile terminally truncated Pdre elements are located at paralogous positions compared with Pgga and the mammalian P-homologous sequences, the data obtained from zebrafish do not support the "source gene hypothesis." They indicate that P-homologous sequences as immobile and subsequently stable components of the genome were generated by "molecular domestication." As a consequence of this explanation, the domestication event of the Phsa/Pgga sequence has occurred by immobilization of an active transposon within a common ancestor before the separation of mammalians and birds about 310 MYA (Nelson 1996).
P-homologous sequences are not the only examples of domesticated transposons. A considerable number of domesticated transposable-element copies now contribute to transcriptionally regulatory elements or to protein-coding regions of cellular genes (Smit 1999, International Human Genome Sequencing Consortium 2001, Jordan et al. 2003). The high number of known transposon-derived domesticated genes provides further evidence that the stable vertebrate P-homologous sequences were recruited as novel genes from their respective host genomes by "molecular domestication" of a former active transposon copy.
![]() |
Supplementary Material |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Acknowledgements |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Footnotes |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Adachi, J., and M. Hasegawa. 1996. Model of amino acid substitution in proteins encoded by mitochondrial DNA. J. Mol. Evol. 42:459468.[ISI][Medline]
Analyse-it for Microsoft Excel. 2003. Version 1.71. Software Ltd., Leeds, UK.
Arnason, U., J. A. Adegoke, K. Bodin, E. W. Born, Y. B. Esa, A. Gullberg, M. Nilsson, R. V. Short, X. Xu, and A. Janke. 2002. Mammalian mitogenomic relationships and the root of the eutherian tree. Proc. Natl. Acad. Sci. USA 99:81518156.
Ausubel, F. A., R. Brent, R. E. Kingston, D. D. Moore, J. G. Seidman, J. A. Smith, and K. Struhl (eds.) 1998. Current protocols in molecular biology. John Wiley and Sons, New York.
Cotton, J. A., and R. D. M. Page. 2002. Going nuclear: gene family evolution and vertebrate phylogeny reconciled. Proc. R. Soc. Lond. B Biol. Sci. 269:15551561.[CrossRef][ISI][Medline]
Daniels S. B., K. R. Peterson, L. D. Strausbaugh, M. G. Kidwell, and A. Chovnick. 1990. Evidence for horizontal transmission of the P transposable element between Drosophila species. Genetics 124:339355.
Dayhoff, M. O., R. M. Schwartz, and B. C. Orcutt. 1978. A model of evolutionary change in proteins. Pp. 345352 in M. O. Dayhoff, ed. Atlas of protein sequence structure, Vol. 5. National Biomedical Research Foundation, Washington, DC.
Farrer, T., A. B. Roller, W. J. Kent, and A. M. Zahler. 2002. Analysis of the role of Caenorhabditis elegans GC-AG introns in regulated splicing. Nucleic Acids Res. 30:33603367.
Felsenstein J. 2004. PHYLIP (phylogeny inference package). Version 3.6b. Distributed by the author, Department of Genome Sciences, University of Washington, Seattle.
Felsenstein J., and G. A. Churchill. 1996. A hidden Markov model approach to variation among sites in rate of evolution. Mol. Biol. Evol. 13:93104.[Abstract]
Gale J. R., M. Blakely, C. M. Hopkins, D. A. Melville, M. W. Wambach, M. Romano, P. R. Katze, and G. Michael. 1998. Regulation of interferon-induced protein kinase PKR: modulation of P58IPK inhibitory function by a novel protein, P52rIBK. Mol. Cell. Biol. 18:859871.
Genetic Analysis Manual: Global Edition IR2 System. 1999. Publication Number 9910-117, LI-COR, Inc. Lincoln, Nebrasha, USA.
Hagemann, S., and W. Pinsker. 2001. Drosophila P transposons in the human genome? Mol. Biol. Evol. 18:19791982.
Hagemann, S., E. Haring, and W. Pinsker. 1996. Repeated horizontal transfer of P transposons between Scaptomyza pallida and Drosophila bifasciata. Genetica 98:4351.[CrossRef][ISI][Medline]
Haring, E., S. Hagemann, and W. Pinsker. 2000. Ancient and recent horizontal invasions of drosophilids by P elements. J. Mol. Evol. 51:577586.[ISI][Medline]
Hasegawa, M., H. Kishino, and K. Yano. 1985. Dating of the human-ape splitting by a molecular clock of mitochondrial DNA. J. Mol. Evol. 22:160174.[ISI][Medline]
International Human Genome Sequencing Consortium.2001. Initial sequencing and analysis of the human genome. Nature 409:860921.[CrossRef][ISI][Medline]
Jones, D. T., W. R. Taylor, and J. M. Thornton. 1992. The rapid generation of mutation data matrices from protein sequences. CABIOS 8:275282.[Medline]
Jordan, I. K., I. B. Rogozin, G. V. Glazko, and E. V. Koonin. 2003. Origin of a substantial fraction of human regulatory sequences from transposable elements. Trends Genet. 19:6872.[CrossRef][ISI][Medline]
Kato, M., A. Miura, J. Bender, S. E. Jacobsen, and T. Kakutani. 2003. Role of CG and non-CG methylation in immobilization of transposons in Arabidopsis. Curr. Biol. 13:421426.[CrossRef][ISI][Medline]
Kidwell, M. G., J. F. Kidwell, and J. A. Sved. 1977. Hybrid dysgenesis in Drosophila melanogaster: a syndrome of aberrant traits including mutation, sterility and male recombination. Genetics 86:813833.
Koenig, M., M. Reichel, R. Marschalek, O. A. Haas, and S. Strehl. 2002. A highly specific and sensitive fluorescence in situ hybridization assay for detection of t(4;11)(q21;q23) and concurrent submicroscopic deletions in acute leukaemias. Brit. J. Haematol. 116:758764.[CrossRef][ISI][Medline]
Lee, S. H., J. B. Clark, and M. G. Kidwell. 1999. A P element-homologous sequence in the house fly, Musca domestica. Insect Mol. Biol. 8:491500.[CrossRef][ISI][Medline]
Marchler-Bauer, A., J. B. Anderson, C. DeWeese-Scott, et al. (24 co-authors).2003. CDD: a curated Entrez database of conserved domain alignments. Nucleic Acids Res. 31:383387.
Miller, W. J., S. Hagemann, E. Reiter, and W. Pinsker. 1992. P-element homologous sequences are tandemly repeated in the genome of Drosophila guanche. Proc. Natl. Acad. Sci. USA 89:40184022.
Miller, W. J., J. F. McDonald, D. Nouaud, and D. Anxolabéhère. 1999. Molecular domesticationmore than a sporadic episode in evolution? Genetica 107:197207.[CrossRef][ISI][Medline]
Misra, S., and D. C. Rio. 1990. Cytotype control of P element transposition: the 66 kD protein is a repressor of transposase activity. Cell 62:269284.[ISI][Medline]
Montgomery, M. K. 2004. RNA interference: historical overview and significance. Methods Mol. Biol. 265:321.[Medline]
Nei, M., and G. V. Glazko. 2002. Estimation of divergence times for a few mammalian and several Primate species. J. Hered. 93:157164.
Nekrutenko A., and W.-H. Li. 2001. Transposable elements are found in a large number of human protein-coding genes. Trends Genet. 17:619621.[CrossRef][ISI][Medline]
Nelson D. R. 1996. Molecular evolution A lecture series by David R. Nelson, placed on the Internet on 21 November, 1996. http://www.icp.ucl.ac.be/opperd/private/m_e_index.html.
Nikaido, M., K. Kawai, Y. Cao, M. Harada, S. Tomita, N. Okada, and M. Hasegawa. 2001. Maximum likelihood analysis of the complete mitochondrial genomes of the Eutherians and a reevaluation of the phylogeny of bats and insectivores. J. Mol. Evol. 53:508516.[CrossRef][ISI][Medline]
Nouaud, D., and D. Anxolabéhère. 1997. P element domestication: a stationary truncated P element may encode a 66-kDa repressor-like protein in the Drosophila montium species subgroup. Mol. Biol. Evol. 14:11321144.[Abstract]
Novina, C. D. and P. A. Sharp. 2004. The RNAi revolution. Nature 430:161164.[CrossRef][ISI][Medline]
O'Hare, K., and G. M. Rubin. 1983. Structures of P transposable elements and their sites of insertion and excision in the Drosophila melanogaster genome. Cell 34:2535.[ISI][Medline]
Oliveira de Carvalho, M., J. C. Silva, and E. L. Loreto.2004. Analyses of P-like transposable element sequences from the genome of Anopheles gambiae. Insect Mol. Biol. 13:5563.[ISI][Medline]
Perkins, H. D., and A. J. Howells. 1992. Genomic sequences with homology to the P element of Drosophila melanogaster occur in the blowfly Lucilia cuprina. Proc. Natl. Acad. Sci. USA 89:1075310757.
Pinsker, W., E. Haring, S. Hagemann, and W. J. Miller. 2001. The evolutionary life history of P transposons: from horizontal invaders to domesticated neogenes. Chromosoma 110:148158.[ISI][Medline]
Posada, D. and K. A. Crandall. 1998. Modeltest: testing the model of DNA substitution. Bioinformatics 17:17818.
Rio, D. C. 1990. Molecular mechanism regulating Drosophila P element transposition. Annu. Rev. Genet. 24:543578.[CrossRef][ISI][Medline]
Roussigne M., C. Cayrol, T. Clouaire, F. Amalric, and J. P. Girard. 2003a. THAP1 is a nuclear proapoptotic factor that links prostate-apoptosis-response-4 (Par-4) to PML nuclear bodies. Oncogene 22:24322442.[CrossRef][ISI][Medline]
Roussigne M., S. Kossida, A. C. Lavigne, T. Clouaire, V. Ecochard, A. Glories, F. Amalric, and J. P. Girard. 2003b. The THAP domain: a novel protein motif with similarity to the DNA-binding domain of P element transposase. Trends Biochem. Sci. 28:6669.[CrossRef][ISI][Medline]
Saitou, N., and M. Nei. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4:14061425.
Sambrook, J., E. F. Fritsch, and T. Maniatis. 1989. Molecular cloninga laboratory manual. 2nd edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
Sarkar, A., R. Rengupta, J. Krzywinski, X. Wang, C. Roth, and F. H. Collins. 2003. P elements are found in the genomes of nematoceran insects of the genus Anopheles. Insect Biochem. Mol. Biol. 33:381387.[CrossRef][ISI][Medline]
Schmidt, H. A., K. Strimmer, M. Vingron, and A. von Haeseler. 2002. TREE-PUZZLE: Maximum likelihood phylogenetic analysis using quartets and parallel computing. Bioinformatics 18:502504.
Silva, J. C., and M. G. Kidwell. 2000. Horizontal transfer and selection in the evolution of P elements. Mol. Biol. Evol. 12:391404.
Smit A. F. A. 1999. Interspersed repeats and other mementos of transposable elements in mammalian genomes. Curr. Opin. Genet. Dev. 9:657663.[CrossRef][ISI][Medline]
Sulston, J. E., and R. Waterston.1998. Toward a complete human genome sequence. Genome Res. 8:10971108.
Tamura, K., and M. Nei. 1993. Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol. Biol. Evol. 10:512526.[Abstract]
Thompson, J. D., T. J. Gibson, F. Plewniak, F. Jeanmougin, and D. G. Higgins. 1997. The ClustalX windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 24:48764882.[CrossRef]
Zhou, Y., E. B. Cambareri, and J. A. Kinsey. 2001. DNA methylation inhibits expression and transposition of the Neurospora Tad retrotransposon. Mol. Genet. Genomics 265:748754.[CrossRef][ISI][Medline]