BACKGROUND
atp-binding cassette  transporters constitute a group of structurally related transmembrane proteins that translocate a vast variety of substrates across cellular membranes and thus perform essential biological roles in prokaryotic and eukaryotic cells  <cit> .
the substrates include lipids, ions, sugars, peptides, vitamins, steroid hormones and xenobiotics such as anticancer drugs.
currently, the human abc transporter family comprises  <dig> members  <cit> .
structurally, functional abc transporters are characterized by two nucleotide binding folds  with conserved walker a and b motifs, signature sequences, and two transmembrane domains , each typically composed of multiple transmembrane segments  <cit> .
the energy required for the transmembrane transport of substrates is provided by hydrolysis of atp at the two nbfs, whereas substrate binding is thought to be determined by the transmembrane and cytosolic domains.
functional abc transporters require the presence of two nbfs and two transmembrane domains.
this dual arrangement is accomplished either by a single polypeptide  or by dimerization of two half-size transporters.
based on sequence similarities, the human abc transporter family has been categorized into seven subfamilies, denoted abc a-g. the human abc a-subfamily presently comprises  <dig> full-size transporters  <cit>  which share common structural features including a large extracellular loop between the first and second transmembrane helix and two conserved 80-amino acid segments adjacent to the nbfs.
to date, four abc a-subfamily genes have been causatively linked to monogenetic lipid trafficking disorders.
these include familial high-density lipoprotein deficiency , neonatal surfactant deficiency , several forms of retinal dystrophies  and two types of hereditary keratinization disorders   <cit> .
recently, a new member of the rodent abc a-transporter subfamily, designated abca <dig>  has been identified which codes for a  <dig> amino acid full-size transporter  <cit> .
the function of abca <dig> is presently unknown, however, its exclusive expression in spermatocytes suggests that this novel transporter may be implicated in lipid transport during spermatogenesis.
in this study, we identified the human ortholog of abca <dig>  we report the surprising finding that the human equivalent of the rodent abca <dig> gene is a transcribed, nonprocessd pseudogene  which forms a unique complex of overlapping genes with the surfactant deficiency gene abca <dig>
RESULTS
bioinformatic prediction of human abca17
homology searches based on the cdna sequences of rat and mouse abca <dig> and of human abca <dig>  respectively, revealed the existence of a homologous gene in the human genome.
the identified gene locus on human chromosome 16p <dig>  exhibits  <dig> %,  <dig> % and  <dig> % identity with the respective cdna sequences of mouse/rat abca <dig> and human abca <dig>  the identification of several est sequences with identities >99%, as retrieved by blat search , provided an additional independent clue for the existence of a human homolog of the rodent abca <dig> gene and it also strongly suggested that it was transcriptionally active.
moreover, homology analyses using the gene prediction program gnomon resulted in the identification of a  <dig> base pair  cdna sequence  denoted "homo sapiens similar to atp-binding cassette, sub-family a member 3; atp-binding cassette 3; abc transporter  <dig> , mrna".
this sequence exhibits 100% identity with the retrieved genomic sequence which is composed of  <dig> exons.
it contains an open reading frame encoding a putative  <dig> amino acid  polypeptide with two nucleotide binding folds and two transmembrane domains, each consisting of three membrane spanning segments and thus predicts a novel member of the abc a-transporter subfamily.
the human homolog of the rodent abca <dig> gene is transcriptionally active but contains numerous mutations
to authenticate the existence of a transcribed human homolog of rodent abca <dig>  we performed rt-pcr using multiple sets of primers specific for the putative abca <dig> coding sequence.
in fact, this approach confirmed that the human homolog of abca <dig> is expressed and sequencing of overlapping pcr fragments revealed a full-length transcript of  <dig>  kb size.
the detailed cdna sequence has been deposited with the ncbi genbank database .
the  <dig>  kb transcript is significantly shorter than that observed for mouse  and rat abca <dig> , respectively, strongly suggesting that the human gene is truncated.
unexpectedly, detailed analysis of the human abca <dig> cdna revealed the presence of numerous mutations including deletions, frameshifts and premature stop codons which preclude its translation into a functional abc transporter protein product.
based on this, we refer to the human homolog of rodent abca <dig> as "abca17p".
an alignment of a hypothetical translation of the abca17p main transcript with the mouse abca <dig> polypeptide sequence is shown in figure  <dig> which details the topology of the abca17p mutations relative to its functional murine counterpart.
the longest open reading frame is located in the center portion of the abca17p cdna sequence and comprises  <dig> bp.
it predicts a hypothetical abc transporter fragment of  <dig> amino acids size which contains three transmembrane spanning segments that corresponds to the n-terminal moiety of the second transmembrane domain of full-size abc transporters .
as expected, the human abca17p nucleotide sequence displays highest identity with mouse  and rat  abca <dig>  these relatively low overall sequence homologies are not surprising because of the presence of several non-homologous gaps within the abca17p sequence.
consistent with this, more detailed segmental blat analysis revealed significantly higher homologies  between the human abca17p gene and the corresponding exons of the mouse abca <dig> gene .
moreover, interspecies comparison of the genomic microenvironments revealed that the human, mouse and rat genes are flanked by homologous genes indicating that the abca17p gene is indeed the human ortholog of the rodent abca <dig> genes.
gene structure of human abca17p and identification of a novel mouse abca <dig> exon 1
the gene structure of the human abca17p gene was determined by blat analysis on the basis of the nucleotide sequence of the identified full-length transcript.
the latter displayed 100% identity with a total of  <dig> adjacent genomic segments  which span an  <dig> kb region on chromosome 16p <dig>  .
of note, systematic database searches and rt-pcr cloning experiments revealed transcripts that originate from an additional first exon  positioned  <dig>  kb upstream of the initially identified exon  <dig>  indicating that the human abca17p gene is in fact composed of  <dig> exons.
the size of the human abca17p gene corresponds to that of other abc a-subfamily members which ranges from  <dig> kb  to  <dig> kb .
however, the fact that functional abc a-transporters are routinely encoded by at least  <dig> exons the presence of only  <dig> exonic segments strongly suggests that abca17p is a truncated gene.
analysis of the exon/intron organization of abca17p demonstrated that exon sizes range from  <dig> to  <dig> bp and introns vary in size between  <dig>  and  <dig>  kb .
all exon-intron boundaries showed the presence of the canonical dinucleotide gt-ag configuration which are diagnostic of splice junctions  <cit> .
surprisingly, close examination of the chromosomal microenvironment of the abca17p locus revealed that it borders on another abc a-transporter gene, the lung surfactant deficiency gene abca <dig>  both genes are arranged in head-to-head orientation on opposite strands and overlap at their 5' ends .
importantly, we found that the abca17p exon 1b is localized between exons  <dig> and  <dig> of the abca <dig> gene and the overlap region  spans  <dig> kb.
thus, the human abca17p/abca <dig> locus represents a unique overlapping complex of a gene and its pseudogene from which both a protein-coding and a non-coding rna are transcribed .
beside the observation of overlapping 5' ends, we found significant overall homology between the human abca17p and abca <dig> genes .
segmental sequence comparison revealed discrete regions of high sequence identity between both genes .
for example, exons 6– <dig> and exons 13– <dig>  respectively, of the abca17p gene exhibit sequence homologies ranging between 70% and 98% with distinct exons of the abca <dig> gene .
moreover, we observed that abca <dig> exon  <dig> is highly homologous to a sequence within intron  <dig> of the abca17p gene.
these sequence homologies in exonic regions together with the chromosomal neighborhood of the abca17p and abca <dig> genes strongly suggest that both genes have originated by duplication of an ancestral gene.
interestingly, cdna sequence comparisons revealed that exon  <dig> of abca17p corresponds to exon  <dig> of the published rodent abca <dig> gene suggesting the existence of an additional 5' exon in the rodent genome.
indeed, similarity searches in est databases led to the identification of an est sequence  which exhibits nearly 100% identity at its 3' end with the 5' portion of the published mouse abca <dig> cdna.
rt-pcr and sequencing experiments confirmed the existence of the novel first exon in the mouse abca <dig> gene .
the novel exon  <dig> is localized  <dig>  kb upstream of the previously published first exon and thus places the mouse abca <dig> gene in closer proximity to its neighbor abca <dig> with an intergenic region of only  <dig>  kb .
of note, the novel exon  <dig> displays only low homology  with abca17p exon 1a and 1b, respectively, indicating that the first exons of the mouse abca <dig> gene and its human ortholog are structurally unrelated.
in the rat genome, the abca <dig> and abca <dig> genes are arranged in the same head-to-head orientation as in the mouse and human genomes.
both genes border on each other but do not overlap and each of the genes encodes a full-size abc transporter.
in addition, we also found evidence for tandem arrangement of the abca <dig> and abca <dig> genes in the dog and cow genomes .
to test whether or not the abca <dig> gene is intact in additional mammalian species, we assembled the putative abca <dig> cdna sequences of dog, cow, chimpanzee and rhesus monkey, respectively, based on available genomic information and sequence identity with the mouse abca <dig> cdna.
using this approach, we identified a  <dig> bp and a  <dig> bp open reading frame in the dog and the cow which potentially encode abca <dig> full-size transporters in either species .
moreover, est database searches demonstrated the existence of several cow est sequences with homologies >98% to the 5' and 3' ends of the predicted cdna strongly suggesting that the cow abca <dig> gene is transcribed.
conversely, analysis of the abca <dig> loci in the chimpanzee and the rhesus monkey genomes revealed numerous sequence alterations including preliminary stop codons, frameshifts, and splice site mutations in multiple exon candidates  consistent with the view that the abca <dig> genes of both of these primate species are indeed pseudogenes.
intriguingly, during our cloning experiments we found evidence for multiple alternatively spliced transcripts of the human abca17p gene.
moreover, combined database searches and cloning experiments revealed the presence of an alternative transcription start site upstream of the one that we initially identified which initiates an alternative exon  <dig> .
the sequence of the alternative exon 1b has been deposited in the ncbi genbank under .
together, these findings predict that up to  <dig> rna variants are transcribed from the human abca17p gene .
our sequencing experiments using pooled cdna derived from  <dig> individuals, also resulted in the identification of two single nucleotide polymorphisms  in the abca17p transcript.
tissue-specific expression of human abca17p
the tissue-specific expression of human abca17p was characterized using multiple tissue northern  blots and the bd™ multiple tissue expression human array  <dig>  containing poly-a+ mrna from various human tissues.
consistent with our rt-pcr cloning results, northern blot analysis demonstrated the existence of an abca17p rna of app.
<dig>  kb size.
this transcript was present in virtually all tissues examined .
highest expression was found in glandular tissue , heart  and fetal tissues  .
in contrast, abca17p rna expression levels in testis and lung were rather low.
the tissue-specific expression pattern of abca17p thus clearly differs from that of its genomic neighbor abca <dig> and also its rodent ortholog abca <dig> which are almost exclusively expressed in the lung and the testis, respectively  <cit> .
discussion
in the present study, we identified the human ortholog of the rodent abc transporter abca <dig>  unexpectedly, we found that this novel abc a-transporter member is a ubiquitously transcribed pseudogene  which structurally overlaps with the gene for the lung surfactant regulator abca <dig>  the existence of a pseudogene homologous to abca <dig>  has recently been proposed in an in silico genomic survey  <cit> .
the authors noted that abca3p <dig> consists of  <dig> exons and borders on the abca <dig> gene at a distance of  <dig> kb.
our results strongly suggest that abca17p and abca3p <dig> are identical, however, they indicate that abca17p/abca3p <dig> consists of a total of  <dig> exons and overlaps with the abca <dig> gene at their common 5' end.
overlapping genes are often functionally or structurally related and genes transcribed from opposite strands of the same genomic locus may, depending on the exact structure of the locus, either regulate each other through antisense-based mechanisms, or may be coordinately regulated due to the common use of a bidirectional promoter  <cit> .
an example for such a functional complex of overlapping genes is the abcg5/abcg <dig> locus.
the abcg <dig> and abcg <dig> genes, which encode two highly homologous half-size abc transporters, are juxtaposed on chromosome 2p <dig> and transcribed from a common bidirectional promoter which drives their coordinate expression.
our finding that the abca17p and abca <dig> genes overlap is thus not without precedence within the human abc transporter gene family.
however, the situation in the abca17p/abca <dig> locus clearly differs from that of the abcg5/abcg <dig> gene complex because abca17p and abca <dig> partially overlap in their transcribed regions and both genes exhibit significant differences in their tissue-specific expression patterns.
the immediate physical proximity of the human abca17p and abca <dig> genes is also observed in the mouse.
systematic analysis of the mouse abca <dig> – abca <dig> gene border resulted in the identification of a novel first exon of the abca <dig> gene, which is located  <dig>  kb upstream of the previously reported exon  <dig>  <cit> .
our findings narrow down the distance between the first exons of the abca <dig> and abca <dig> genes to a region of only  <dig> kb, however, we did not find evidence that both transcription units overlap.
consistent with this, the abca17p expression pattern differs considerably from that of its rodent ortholog abca <dig> which is exclusively expressed in the testis  <cit> .
it is thus likely that the overlap between the 5' ends of the abca17p and abca <dig> genes, which is absent from the mouse abca17/abca <dig> locus, constitutes the physical basis for the observed disparate tissue-specific expression of human abca17p and mouse abca <dig>
the abca17p/abca <dig> locus is unique because it is the first demonstration in the human genome that a pseudogene and its progenitor gene overlap.
to our knowledge, this is also the first example of a pseudogene/gene pair that is transcribed from a common 5' end.
the significant homology between the abca17p gene and its neighboring progenitor abca <dig> and their colocalization in the genome strongly suggest that both genes arose by duplication of an ancestor gene.
the fact that abca <dig> and abca <dig> are also neighbors in the rodent genome and the genomes of the dog and the cow, respectively, indicates that the gene duplication arose before the boreoeutherian radiation which has been dated to app.
<dig> myr  <cit> .
moreover, our combined analyses of the genomes of the above species and those of the chimpanzee and the rhesus monkey suggest that the pseudogenization of the abca <dig> locus occurred after the divergence of rodents and primates.
given the fact that mutations in the abca <dig> gene cause neonatal surfactant deficiency  <cit> , our identification of the overlapping abca17p/abca <dig> locus confirms the observation that overlapping genes show a striking association with disease genes  <cit> .
the distance between pseudogenes and their progenitor genes is inversely correlated with the likelihood for the transfer of mutations from a pseudogene to its functional gene by gene conversion  <cit> .
for example, it has been shown that deleterious mutations from abcc <dig> pseudogenes are transferred to their parent gene abcc <dig>  <cit> .
in contrast to the two known abcc <dig> pseudogenes, which are located  <dig>  mb telomeric and  <dig>  mb centromeric of abcc <dig>  respectively, the distance between abca17p and its functional gene abca <dig> is zero.
thus, there is a high probability that abca17p serves as a repository for mutations in the abca <dig> gene.
however, detailed sequence comparison between the known mutated exons in abca <dig> and their corresponding exons in abca17p revealed that all nucleotide positions in the abca17p gene that correspond to mutation sites in abca <dig> represent wildtype sequences.
moreover, in silico searches for gene conversion events in the abca17p exons 6– <dig> and 13– <dig>  which share highest homology with abca <dig> exons, utilizing the program geneconv did not yield statistically significant results .
together, these findings do not support the view that sequences of abca17p may have contributed mutations to abca <dig>
importantly, we also found that abca17p and abca <dig> genes share discrete regions of high homology.
in particular, exons  <dig> and  <dig> of the abca17p gene exhibit up to 98% sequence identity with the abca <dig> exons  <dig> and  <dig>  respectively.
the existence of these highly homologous dna segments in both genes has immediate practical relevance because accidental amplification of abca17p sequences may provide erroneous results upon mutation analysis of the abca <dig> gene.
because abca17p is transcriptionally active in a broad spectrum of human tissues including the lung, the major production site of abca <dig>  it also needs to be taken into account that abca17p transcripts may potentially interfere with abca <dig> mrna expression analyses due to cross-hybridization of abca <dig> specific probes or rt-pcr primers with homologous pseudogene sequences.
although recent computational approaches estimate that the number of pseudogenes in the human genome ranges between  <dig>  and  <dig>   <cit> , only little is known about their biological significance.
in particular, there is a great paucity of information on transcriptionally active pseudogenes.
considering that the most comprehensive study thus far focussing on transcribed pseudogenes suggests that up to 20% of the pseudogenes on chromosome  <dig> may indeed be transcriptionally active  <cit> , it is tempting to speculate that transcribed pseudogenes may serve defined biological functions rather than represent mere genomic fossils.
this view is convincingly supported by a recent landmark paper that provided evidence for a specific function of a transcribed pseudogene by demonstrating that the expressed pseudogene makorin1-p <dig> regulates the stability of the mrna of its progenitor gene makorin <dig>  <cit> .
another mechanism of specific pseudogene-gene interference has been identified for the antisense pseudogene anti-nos whose transcript forms stable rna-rna complexes with the mrna of nnos and thus inhibits protein expression of its parent gene  <cit> .
at this point, the exact function of the abca17p gene is unclear.
however, abca17p stands out among the known transcribed pseudogenes in that it shares its 5' end with its functional counterpart.
due to this unique genomic architecture the putative abca17p promoter is part of the abca <dig> gene and vice versa.
moreover, the primary transcripts derived from both genes have a  <dig>  kb overlap.
it is thus conceivable that abca17p and abca <dig> may mutually impact their promoter activities and the elongation rates of their primary transcripts due to steric interference of their transcription machineries.
such a scenario is supported by the largely complementary expression patterns of both genes in human tissues.
more work is required to determine whether and to which degree abca17p is involved in the control of the disease gene abca <dig>
CONCLUSIONS
in this study, we identified the primary structure of the human ortholog of the recently cloned murine abc transporter abca <dig> and demonstrate that, unlike its rodent counterpart, it is a ubiquitously expressed pseudogene.
surprisingly, we found that abca17p overlaps with the gene for the lung surfactant regulator abca <dig> and sequence homology analyses strongly suggest that abca <dig> is the progenitor gene of abca17p.
to our knowledge, this is the first example of the co-expression of a pseudogene and its progenitor gene from a common overlapping dna region in the human genome.
these unique genomic features suggest potential interdependencies between the regulation of abca17p and the disease gene abca <dig>

 atp-binding cassette transporters constitute a group of structurally related transmembrane proteins that translocate a vast variety of substrates across cellular membranes and thus perform essential biological roles in prokaryotic and eukaryotic cells. the substrates include lipids, ions, sugars, peptides, vitamins, steroid hormones and xenobiotics such as anticancer drugs. currently, the human abc transporter family comprises members. structurally, functional abc transporters are characterized by two nucleotide binding folds with conserved walker a and b motifs, signature sequences, and two transmembrane domains, each typically composed of multiple transmembrane segments. the energy required for the transmembrane transport of substrates is provided by hydrolysis of atp at the two nbfs, whereas substrate binding is thought to be determined by the transmembrane and cytosolic domains. functional abc transporters require the presence of two nbfs and two transmembrane domains. this dual arrangement is accomplished either by a single polypeptide or by dimerization of two half-size transporters. based on sequence similarities, the human abc transporter family has been categorized into seven subfamilies, denoted abc a-g. the human abc a-subfamily presently comprises full-size transporters which share common structural features including a large extracellular loop between the first and second transmembrane helix and two conserved 80-amino acid segments adjacent to the nbfs. to date, four abc a-subfamily genes have been causatively linked to monogenetic lipid trafficking disorders. these include familial high-density lipoprotein deficiency, neonatal surfactant deficiency, several forms of retinal dystrophies and two types of hereditary keratinization disorders. recently, a new member of the rodent abc a-transporter subfamily, designated abca has been identified which codes for a amino acid full-size transporter. the function of abca is presently unknown, however, its exclusive expression in spermatocytes suggests that this novel transporter may be implicated in lipid transport during spermatogenesis. in this study, we identified the human ortholog of abca we report the surprising finding that the human equivalent of the rodent abca gene is a transcribed, nonprocessd pseudogene which forms a unique complex of overlapping genes with the surfactant deficiency gene abca  bioinformatic prediction of human abca17 homology searches based on the cdna sequences of rat and mouse abca and of human abca respectively, revealed the existence of a homologous gene in the human genome. the identified gene locus on human chromosome 16p exhibits %, % and % identity with the respective cdna sequences of mouse/rat abca and human abca the identification of several est sequences with identities >99%, as retrieved by blat search, provided an additional independent clue for the existence of a human homolog of the rodent abca gene and it also strongly suggested that it was transcriptionally active. moreover, homology analyses using the gene prediction program gnomon resulted in the identification of a base pair cdna sequence denoted "homo sapiens similar to atp-binding cassette, sub-family a member 3; atp-binding cassette 3; abc transporter, mrna". this sequence exhibits 100% identity with the retrieved genomic sequence which is composed of exons. it contains an open reading frame encoding a putative amino acid polypeptide with two nucleotide binding folds and two transmembrane domains, each consisting of three membrane spanning segments and thus predicts a novel member of the abc a-transporter subfamily. the human homolog of the rodent abca gene is transcriptionally active but contains numerous mutations to authenticate the existence of a transcribed human homolog of rodent abca we performed rt-pcr using multiple sets of primers specific for the putative abca coding sequence. in fact, this approach confirmed that the human homolog of abca is expressed and sequencing of overlapping pcr fragments revealed a full-length transcript of kb size. the detailed cdna sequence has been deposited with the ncbi genbank database. the kb transcript is significantly shorter than that observed for mouse and rat abca, respectively, strongly suggesting that the human gene is truncated. unexpectedly, detailed analysis of the human abca cdna revealed the presence of numerous mutations including deletions, frameshifts and premature stop codons which preclude its translation into a functional abc transporter protein product. based on this, we refer to the human homolog of rodent abca as "abca17p". an alignment of a hypothetical translation of the abca17p main transcript with the mouse abca polypeptide sequence is shown in figure which details the topology of the abca17p mutations relative to its functional murine counterpart. the longest open reading frame is located in the center portion of the abca17p cdna sequence and comprises bp. it predicts a hypothetical abc transporter fragment of amino acids size which contains three transmembrane spanning segments that corresponds to the n-terminal moiety of the second transmembrane domain of full-size abc transporters. as expected, the human abca17p nucleotide sequence displays highest identity with mouse and rat abca these relatively low overall sequence homologies are not surprising because of the presence of several non-homologous gaps within the abca17p sequence. consistent with this, more detailed segmental blat analysis revealed significantly higher homologies between the human abca17p gene and the corresponding exons of the mouse abca gene. moreover, interspecies comparison of the genomic microenvironments revealed that the human, mouse and rat genes are flanked by homologous genes indicating that the abca17p gene is indeed the human ortholog of the rodent abca genes. gene structure of human abca17p and identification of a novel mouse abca exon 1 the gene structure of the human abca17p gene was determined by blat analysis on the basis of the nucleotide sequence of the identified full-length transcript. the latter displayed 100% identity with a total of adjacent genomic segments which span an kb region on chromosome 16p. of note, systematic database searches and rt-pcr cloning experiments revealed transcripts that originate from an additional first exon positioned kb upstream of the initially identified exon indicating that the human abca17p gene is in fact composed of exons. the size of the human abca17p gene corresponds to that of other abc a-subfamily members which ranges from kb to kb. however, the fact that functional abc a-transporters are routinely encoded by at least exons the presence of only exonic segments strongly suggests that abca17p is a truncated gene. analysis of the exon/intron organization of abca17p demonstrated that exon sizes range from to bp and introns vary in size between and kb. all exon-intron boundaries showed the presence of the canonical dinucleotide gt-ag configuration which are diagnostic of splice junctions. surprisingly, close examination of the chromosomal microenvironment of the abca17p locus revealed that it borders on another abc a-transporter gene, the lung surfactant deficiency gene abca both genes are arranged in head-to-head orientation on opposite strands and overlap at their 5' ends. importantly, we found that the abca17p exon 1b is localized between exons and of the abca gene and the overlap region spans kb. thus, the human abca17p/abca locus represents a unique overlapping complex of a gene and its pseudogene from which both a protein-coding and a non-coding rna are transcribed. beside the observation of overlapping 5' ends, we found significant overall homology between the human abca17p and abca genes. segmental sequence comparison revealed discrete regions of high sequence identity between both genes. for example, exons 6– and exons 13– respectively, of the abca17p gene exhibit sequence homologies ranging between 70% and 98% with distinct exons of the abca gene. moreover, we observed that abca exon is highly homologous to a sequence within intron of the abca17p gene. these sequence homologies in exonic regions together with the chromosomal neighborhood of the abca17p and abca genes strongly suggest that both genes have originated by duplication of an ancestral gene. interestingly, cdna sequence comparisons revealed that exon of abca17p corresponds to exon of the published rodent abca gene suggesting the existence of an additional 5' exon in the rodent genome. indeed, similarity searches in est databases led to the identification of an est sequence which exhibits nearly 100% identity at its 3' end with the 5' portion of the published mouse abca cdna. rt-pcr and sequencing experiments confirmed the existence of the novel first exon in the mouse abca gene. the novel exon is localized kb upstream of the previously published first exon and thus places the mouse abca gene in closer proximity to its neighbor abca with an intergenic region of only kb. of note, the novel exon displays only low homology with abca17p exon 1a and 1b, respectively, indicating that the first exons of the mouse abca gene and its human ortholog are structurally unrelated. in the rat genome, the abca and abca genes are arranged in the same head-to-head orientation as in the mouse and human genomes. both genes border on each other but do not overlap and each of the genes encodes a full-size abc transporter. in addition, we also found evidence for tandem arrangement of the abca and abca genes in the dog and cow genomes. to test whether or not the abca gene is intact in additional mammalian species, we assembled the putative abca cdna sequences of dog, cow, chimpanzee and rhesus monkey, respectively, based on available genomic information and sequence identity with the mouse abca cdna. using this approach, we identified a bp and a bp open reading frame in the dog and the cow which potentially encode abca full-size transporters in either species. moreover, est database searches demonstrated the existence of several cow est sequences with homologies >98% to the 5' and 3' ends of the predicted cdna strongly suggesting that the cow abca gene is transcribed. conversely, analysis of the abca loci in the chimpanzee and the rhesus monkey genomes revealed numerous sequence alterations including preliminary stop codons, frameshifts, and splice site mutations in multiple exon candidates consistent with the view that the abca genes of both of these primate species are indeed pseudogenes. intriguingly, during our cloning experiments we found evidence for multiple alternatively spliced transcripts of the human abca17p gene. moreover, combined database searches and cloning experiments revealed the presence of an alternative transcription start site upstream of the one that we initially identified which initiates an alternative exon. the sequence of the alternative exon 1b has been deposited in the ncbi genbank under. together, these findings predict that up to rna variants are transcribed from the human abca17p gene. our sequencing experiments using pooled cdna derived from individuals, also resulted in the identification of two single nucleotide polymorphisms in the abca17p transcript. tissue-specific expression of human abca17p the tissue-specific expression of human abca17p was characterized using multiple tissue northern blots and the bd™ multiple tissue expression human array containing poly-a+ mrna from various human tissues. consistent with our rt-pcr cloning results, northern blot analysis demonstrated the existence of an abca17p rna of app. kb size. this transcript was present in virtually all tissues examined. highest expression was found in glandular tissue, heart and fetal tissues. in contrast, abca17p rna expression levels in testis and lung were rather low. the tissue-specific expression pattern of abca17p thus clearly differs from that of its genomic neighbor abca and also its rodent ortholog abca which are almost exclusively expressed in the lung and the testis, respectively. discussion in the present study, we identified the human ortholog of the rodent abc transporter abca unexpectedly, we found that this novel abc a-transporter member is a ubiquitously transcribed pseudogene which structurally overlaps with the gene for the lung surfactant regulator abca the existence of a pseudogene homologous to abca has recently been proposed in an in silico genomic survey. the authors noted that abca3p consists of exons and borders on the abca gene at a distance of kb. our results strongly suggest that abca17p and abca3p are identical, however, they indicate that abca17p/abca3p consists of a total of exons and overlaps with the abca gene at their common 5' end. overlapping genes are often functionally or structurally related and genes transcribed from opposite strands of the same genomic locus may, depending on the exact structure of the locus, either regulate each other through antisense-based mechanisms, or may be coordinately regulated due to the common use of a bidirectional promoter. an example for such a functional complex of overlapping genes is the abcg5/abcg locus. the abcg and abcg genes, which encode two highly homologous half-size abc transporters, are juxtaposed on chromosome 2p and transcribed from a common bidirectional promoter which drives their coordinate expression. our finding that the abca17p and abca genes overlap is thus not without precedence within the human abc transporter gene family. however, the situation in the abca17p/abca locus clearly differs from that of the abcg5/abcg gene complex because abca17p and abca partially overlap in their transcribed regions and both genes exhibit significant differences in their tissue-specific expression patterns. the immediate physical proximity of the human abca17p and abca genes is also observed in the mouse. systematic analysis of the mouse abca – abca gene border resulted in the identification of a novel first exon of the abca gene, which is located kb upstream of the previously reported exon. our findings narrow down the distance between the first exons of the abca and abca genes to a region of only kb, however, we did not find evidence that both transcription units overlap. consistent with this, the abca17p expression pattern differs considerably from that of its rodent ortholog abca which is exclusively expressed in the testis. it is thus likely that the overlap between the 5' ends of the abca17p and abca genes, which is absent from the mouse abca17/abca locus, constitutes the physical basis for the observed disparate tissue-specific expression of human abca17p and mouse abca the abca17p/abca locus is unique because it is the first demonstration in the human genome that a pseudogene and its progenitor gene overlap. to our knowledge, this is also the first example of a pseudogene/gene pair that is transcribed from a common 5' end. the significant homology between the abca17p gene and its neighboring progenitor abca and their colocalization in the genome strongly suggest that both genes arose by duplication of an ancestor gene. the fact that abca and abca are also neighbors in the rodent genome and the genomes of the dog and the cow, respectively, indicates that the gene duplication arose before the boreoeutherian radiation which has been dated to app. myr. moreover, our combined analyses of the genomes of the above species and those of the chimpanzee and the rhesus monkey suggest that the pseudogenization of the abca locus occurred after the divergence of rodents and primates. given the fact that mutations in the abca gene cause neonatal surfactant deficiency, our identification of the overlapping abca17p/abca locus confirms the observation that overlapping genes show a striking association with disease genes. the distance between pseudogenes and their progenitor genes is inversely correlated with the likelihood for the transfer of mutations from a pseudogene to its functional gene by gene conversion. for example, it has been shown that deleterious mutations from abcc pseudogenes are transferred to their parent gene abcc. in contrast to the two known abcc pseudogenes, which are located mb telomeric and mb centromeric of abcc respectively, the distance between abca17p and its functional gene abca is zero. thus, there is a high probability that abca17p serves as a repository for mutations in the abca gene. however, detailed sequence comparison between the known mutated exons in abca and their corresponding exons in abca17p revealed that all nucleotide positions in the abca17p gene that correspond to mutation sites in abca represent wildtype sequences. moreover, in silico searches for gene conversion events in the abca17p exons 6– and 13– which share highest homology with abca exons, utilizing the program geneconv did not yield statistically significant results. together, these findings do not support the view that sequences of abca17p may have contributed mutations to abca importantly, we also found that abca17p and abca genes share discrete regions of high homology. in particular, exons and of the abca17p gene exhibit up to 98% sequence identity with the abca exons and respectively. the existence of these highly homologous dna segments in both genes has immediate practical relevance because accidental amplification of abca17p sequences may provide erroneous results upon mutation analysis of the abca gene. because abca17p is transcriptionally active in a broad spectrum of human tissues including the lung, the major production site of abca it also needs to be taken into account that abca17p transcripts may potentially interfere with abca mrna expression analyses due to cross-hybridization of abca specific probes or rt-pcr primers with homologous pseudogene sequences. although recent computational approaches estimate that the number of pseudogenes in the human genome ranges between and, only little is known about their biological significance. in particular, there is a great paucity of information on transcriptionally active pseudogenes. considering that the most comprehensive study thus far focussing on transcribed pseudogenes suggests that up to 20% of the pseudogenes on chromosome may indeed be transcriptionally active, it is tempting to speculate that transcribed pseudogenes may serve defined biological functions rather than represent mere genomic fossils. this view is convincingly supported by a recent landmark paper that provided evidence for a specific function of a transcribed pseudogene by demonstrating that the expressed pseudogene makorin1-p regulates the stability of the mrna of its progenitor gene makorin. another mechanism of specific pseudogene-gene interference has been identified for the antisense pseudogene anti-nos whose transcript forms stable rna-rna complexes with the mrna of nnos and thus inhibits protein expression of its parent gene. at this point, the exact function of the abca17p gene is unclear. however, abca17p stands out among the known transcribed pseudogenes in that it shares its 5' end with its functional counterpart. due to this unique genomic architecture the putative abca17p promoter is part of the abca gene and vice versa. moreover, the primary transcripts derived from both genes have a kb overlap. it is thus conceivable that abca17p and abca may mutually impact their promoter activities and the elongation rates of their primary transcripts due to steric interference of their transcription machineries. such a scenario is supported by the largely complementary expression patterns of both genes in human tissues. more work is required to determine whether and to which degree abca17p is involved in the control of the disease gene abca  in this study, we identified the primary structure of the human ortholog of the recently cloned murine abc transporter abca and demonstrate that, unlike its rodent counterpart, it is a ubiquitously expressed pseudogene. surprisingly, we found that abca17p overlaps with the gene for the lung surfactant regulator abca and sequence homology analyses strongly suggest that abca is the progenitor gene of abca17p. to our knowledge, this is the first example of the co-expression of a pseudogene and its progenitor gene from a common overlapping dna region in the human genome. these unique genomic features suggest potential interdependencies between the regulation of abca17p and the disease gene abca 
this genomic colocalization and the observation that the abca17p and abca genes share significant homologies in several exons suggest that both genes have evolved by gene duplication. our results demonstrate that abca17p and abca form a complex of overlapping genes in the human genome from which both non-coding and protein-coding abc a-transporter rnas are expressed. genomic analysis revealed that abca17p borders on another abc a-subfamily transporter – the lung surfactant deficiency gene abca surprisingly, we found that both genes overlap at their first exons and are transcribed from opposite strands. in this study, we report the identification and characterization of the human ortholog of rodent abca the new abc transporter gene exhibits striking nucleotide sequence homology with the recently cloned mouse and rat abca, respectively, and is located in the syntenic region of mouse abca indicating that it represents the human ortholog of rodent abca however, unlike in the mouse, the full-length abca transcript contains numerous mutations that preclude its translation into a bona fide abc transporter protein strongly suggesting that the human abca gene is a transcribed pseudogene. the fact that both genes overlap at their 5' ends suggests interdependencies in their regulation and may have important implications for the functional analysis of the disease gene abca moreover, this is the first demonstration of the expression of a pseudogene and its parent gene from a common overlapping dna region in the human genome.
 during the past years, we and others discovered a series of human atp-binding cassette transporters, now referred to as abc a-subfamily transporters. recently, a novel testis-specific abc a transporter, abca has been cloned in rodent. in this study, we report the identification and characterization of the human ortholog of rodent abca  the novel human abc a-transporter gene on chromosome 16p is ubiquitously expressed with highest expression in glandular tissues and the heart. the new abc transporter gene exhibits striking nucleotide sequence homology with the recently cloned mouse and rat abca, respectively, and is located in the syntenic region of mouse abca indicating that it represents the human ortholog of rodent abca however, unlike in the mouse, the full-length abca transcript contains numerous mutations that preclude its translation into a bona fide abc transporter protein strongly suggesting that the human abca gene is a transcribed pseudogene. we identified numerous alternative abca17p splice variants which are transcribed from two distinct transcription initiation sites. genomic analysis revealed that abca17p borders on another abc a-subfamily transporter – the lung surfactant deficiency gene abca surprisingly, we found that both genes overlap at their first exons and are transcribed from opposite strands. this genomic colocalization and the observation that the abca17p and abca genes share significant homologies in several exons suggest that both genes have evolved by gene duplication.  our results demonstrate that abca17p and abca form a complex of overlapping genes in the human genome from which both non-coding and protein-coding abc a-transporter rnas are expressed. the fact that both genes overlap at their 5' ends suggests interdependencies in their regulation and may have important implications for the functional analysis of the disease gene abca moreover, this is the first demonstration of the expression of a pseudogene and its parent gene from a common overlapping dna region in the human genome. 