Genomic Organization Analysis Research Articles

Canine PKD1 Is a Single-Copy Gene: Genomic Organization and Comparative Analysis

Most cases of autosomal dominant polycystic kidney disease are caused by mutations in the gene PKD1, encoding polycystin-1. To gain insight into the role of polycystin-1 in tubulogenesis and cystogenesis using the well-characterized canine kidney epithelial cell line MDCK, we have now cloned and characterized the exon/intron structure of the canine gene PKD1. FISH analysis showed that the dog genome lacks the multiple PKD1 homologs present in human. Intron 21 of dog PKD1 lacked the polypyrimidine tract characteristic of the human gene, whereas pyrimidine-rich elements were identified in canine intron 30. Canine polycystin-1 showed a higher degree of homology with the human counterpart and lower homology with mouse and rat. A striking degree of conservation (97% identity) was determined for the leucine-rich repeat domain between dog and human. Also, the homology analysis indicated that 4 of 16 Ig-like repeats (IgIII, IgVII, IgX, and IgXV) are likely to be functionally significant. This is particularly important in light of our recent findings demonstrating that Iglike domains form strong homophilic interactions and can mediate cell–cell adhesion. These data enable detailed analysis of the role of polycystin-1 in cystogenesis and tubulogenesis using the canine MDCK cell line.

Identification of a new small GTP-binding protein gene OsRab5a, genomic organization, and expression pattern analysis during nitrate supply and early nutrient starvation in rice ( Oryza sativa L .) root

A new nitrate-induced gene OsRab5a in rice was isolated using suppression subtractive hybridization (SSH) method. OsRab5a has 1125 nucleotides with an open reading frame of 612 nucleotides encoding a small GTP-binding protein of Rab family. OsRab5a had a higher expression in root, weak expression in shoot, flower and grain, and visually not in stem and leaf. Southern and mapping analysis indicated that OsRab5a would be a single copy gene at locus of 108.8 cM of chromosome 12. Comparison of genomic organization and tissue distribution showed well-conserved genomic organizations and similar expression patterns between OsRab5a gene in rice and Rha1 gene in Arabidopsis during evolution. OsRab5a was quickly induced by nitrate in a split-root treatment, also up-regulated at early nitrogen and phosphorus starvation in root. Acceleration of root growth was observed under nitrate supply, early nitrogen and phosphorous starvation, which was consistent with the expression pattern of OsRab5a. These results suggest that OsRab5a may function in nutrient-regulated root growth.

The Human Monocarboxylate Transporter, MCT1: Genomic Organization and Promoter Analysis

Uptake of butyrate across the colonocyte luminal membrane is mediated by the monocarboxylate transporter isoform 1 (MCT1). We have demonstrated previously that expression of human colonic MCT1 is responsive to butyrate, and that this involves the dual control of MCT1 gene transcription and stability of the MCT1 transcript. Here we describe the structural organization of the human MCT1 gene, and report the isolation and characterization of the MCT1 gene promoter. The MCT1 gene spans approximately 44 kb, and is organized as 5 exons intervened by 4 introns. The first of these introns is located in the 5′-UTR-encoding DNA, spans >26 kb, and thus accounts for approximately 60% of the entire transcription unit. Analysis of a 1.5 kb fragment of the MCT1 5′-flanking region, shows an absence of the classical TATA-Box motif. However, the region contains potential binding sites for a variety of transcription factors with known association with butyrate's action in the colon. In transient transfections the 5′-flanking region drives high-level expression of a luciferase reporter-gene in cells that endogenously express MCT1. Deletion analyses indicate that the cis-acting elements necessary for basal transcription of MCT1 are contained within the −70/+213 proximal sequence of the promoter.

Genomic organization, transcript variants and comparative analysis of the human nucleoporin 155 ( NUP155) gene

Nucleoporin 155 ( Nup155) is a major component of the nuclear pore complex (NPC) involved in cellular nucleo-cytoplasmic transport. We have acquired the complete sequence and interpreted the genomic organization of the Nup155 orthologos from human ( Homo sapiens) and pufferfish ( Fugu rubripes), which are approximately 80 and 8 kb in length, respectively. The human gene is ubiquitously expressed in many tissues analyzed and has two major transcript variants, resulted from an alternative usage of the 5′ cryptic or consensus splice donor in intron 1 and two polyadenylation signals. We have also cloned DNA complementary to RNAs of the Nup155 orthologs from Fugu and mouse. Comparative analysis of the Nup155 orthologs in many species, including H. sapiens, Mus musculus, Rattus norvegicus, F. rubripes, Arabidopsis thaliana, Drosophila melanogaster, and Saccharomyces cerevisiae, has revealed two paralogs in S. cerevisiae but only a single gene with increasing number of introns in more complex organisms. The amino acid sequences of the Nup155 orthologos are highly conserved in the evolution of eukaryotes. Different gene orders in the human and Fugu genomic regions harboring the Nup155 orthologs advocate cautious interpretation of synteny in comparative genomic analysis even within the vertebrate lineage.

Genome organization and phylogenetic tree analysis of Garlic virus E, a new member of genus Allexivirus

The complete sequence of an Allexivirus isolated from garlic plants in Yuhang City, Zhejiang Province, China had been determined. The single-strand, positive RNA genome was 8451 nucleotides in length excluding poly(A) tail. The genome organization of this virus was similar to that of the other Allexiviruses but only with 62.8%–64.8% nucleotide acid identities. The amino acid sequences of proteins encoded by ORF1-6 shared 67.6%–78.5%, 55.4%–66.2%, 56.7%–66.4%,40.3%–55.6%,66.3%–79.7%and 52.2%–68.8% identities with those of the others respectively. The homology range between it and the other Allexiviruses was similar to that between the other distinct species in this genus. A more comprehensive comparison using all available CP amino acid sequences showed that it shared only 63.9%–79.8% amino acids identical with the others. Therefore, it had been considered as a new member of the genus, named as garlic virus E (GarV-E). Phylogenetic analysis confirmed GarV-E as a distinct member and the correct names and classification of some members of genus Allexivirus were also discussed.

Complete nucleotide sequence, genome organization and phylogenic analysis of the canine calicivirus.

The complete genomic sequence of canine calicivirus (CaCV) isolated from feces of a dog with diarrhea was determined. The CaCV genome, a positive-sense single-stranded RNA, contained 8513 nucleotides excluding the poly(A) tail and was longer than that of any other calicivirus strain with a completely known sequence. There were three open reading frames (ORF1, nt 12-5801; ORF2, nt 5805-7880; and ORF3, nt 7877-8278). ORF1 encoded a polyprotein (calculated Mr of 214,802) which had the conserved motifs of non-structural proteins of other caliciviruses and picornaviruses. Regions containing characteristic motifs in the non-structural polyprotein of CaCV showed highest similarity with those of the species Feline calicivirus and Vesicular exanthema of swine virus in the genus Vesivirus. Phylogenic analysis indicated that CaCV formed a distinct branch within the genus. Our results strongly suggested that CaCV is a new species in the genus Vesivirus.

Cloning and characterization of a cDNA encoding a cobalamin-independent methionine synthase from potato (Solanum tuberosum L.).

A potato cDNA clone, StMS1, that encodes a methionine synthase was isolated. This protein was identified on the basis of both structural and functional evidence. The predicted sequence of the protein encoded by StMS1 shows a high degree of similarity to methionine synthases from other organisms and the expression of StMS1 in bacterial mutant strains restored the mutant's ability to synthesize methionine. Genomic organization and expression analyses suggest that StMS1 is a low-copy gene and is differentially expressed in potato organs. StMS1 expression was found in all tissues, but at elevated levels in flowers, basal levels in sink and source leaves, roots and stolons, and low levels in stems and tubers. RNA expression data were confirmed by western blot analysis except that the protein content in leaves was less than expected from the RNA data. Western blot analysis of subcellular fractions revealed that the protein is located in the cytosol. However, the changing pattern of gene expression during the day/night period implied a light-dependent control of MS transcription normally seen for enzymes localized in plastids. The expression of MS was shown to be light-inducible with its highest expression at midday. These RNA data were not confirmed at the protein level since protein content levels remained constant over the whole day. Feeding experiments of detached leaves revealed that sucrose or sucrose-derived products are responsible for StMS1 induction. This induction can be blocked by treatment with DCMU during the light period. Western analysis revealed that the amount of StMS1 is not affected by either treatment. This experiment confirmed the presence of a day/night rhythm. Methionine synthase expression is regulated by photoassimilates but this seems not to detectably alter protein levels.

Identification, genomic organization, chromosomal mapping and mutation analysis of the human INV gene, the ortholog of a murine gene implicated in left-right axis development and biliary atresia

Determination of left-right axis is a precocious embryonic event, and all phenotypic anomalies resulting from disruption of the normal lateralization process are collectively referred to as the lateralization defect. A transgenic mouse with lateralization defect and hepatic, kidney, and pancreatic anomalies has resulted from disruption of the inv gene by insertion of a transgene. The human ortholog is thus a good candidate for lateralization defect in humans, in particular in cases with associated hepatic anomalies. Here, we have identified, mapped, and characterized the INV human gene and screened a series of heterotaxic patients (with or without biliary anomalies) for mutation in this gene. In a German family of Turkish origin, we have found that all available affected and unaffected individuals are heterozygous for a mutation in the splicing donor site of intron 12 in the INV gene resulting in two different aberrant splicing isoforms. This can be explained either by a randomization of lateralization defects or, as suggested earlier, di- or trigenic inheritance, although we have been unable to detect, in this family, a mutation in genes known to be involved in the human lateralization defect ( LEFTY1, LEFTY2, ACVR2B, NODAL, ZIC3, and CFC1). In contrast to the mouse, the affected individuals have no biliary anomalies, and the absence of mutation in a series of seven cases with lateralization defect and biliary anomalies demonstrates that INV is not frequently involved in such a phenotype in humans.

Genomic organization and promoter analysis of mouse follistatin-related gene (FLRG)

Follistatin (FS) is well characterized as an activin-binding protein. Recently, a novel follistatin-like protein called follistatin-related gene (FLRG) that has a similar domain organization to that of follistatin has been identified. Like follistatins, FLRG binds activins and bone morphogenetic proteins (BMPs). To study the regulation of FLRG expression, we have analyzed the genomic organization and promoter of the mouse FLRG gene. The mouse FLRG gene consists of five exons, and each encodes discrete functional regions. The overall genomic structure of FLRG is similar to that of FS except that the FLRG gene is missing one exon that codes a third FS domain found in FS. The promoter that covers 2.5 kbp and is linked to a luciferase reporter construct is active in human cervical carcinoma HeLa cells as well as in human embryonic kidney (HEK293) cells. Deletion analysis of the promoter regions indicates that a proximal 550 base pairs are enough for basal FLRG promoter activity in the cell lines. FLRG promoter activity is significantly augmented by phorbol 12-myristate 13-acetate (PMA) treatment, but not by cAMP stimulation. By contrast, FS promoter is activatable either by cAMP or PMA. Thus, although FS and FLRG are structurally and functionally related, their modes of regulation by external stimuli are different.

Genomic organization, chromosomal localization, and promoter analysis of the mouse Mail gene.

The Mail (molecule possessing ankyrin repeats induced by lipopolysaccharide) protein is a member of the IkappaB family. It has six ankyrin repeats that are conserved in other IkappaB proteins, such as IkappaB-alpha and Bcl-3. Mail mRNA expression is induced rapidly following lipopolysaccharide (LPS) injection, most notably in the spleen, lung, and lymph nodes of mice, where immune cells, such as lymphocytes and macrophages, are abundant. In this study, we cloned and characterized the Mail gene. The isolated genomic clones span approximately 30 kb and encompass the entire gene. Comparisons with Mail cDNA revealed that the Mail gene consists of 14 exons. Several splice junctions encoding ankyrin repeats are conserved among Mail and other IkappaB family genes. Southern hybridization showed that Mail is a single-copy gene. Using fluorescence in situ hybridization analysis, mouse and rat Mail genes were mapped to Chromosome (Chr) 16C1.2-C1.3 and Chr 11q21.1, respectively. Primer extension determined the transcription start site of Mail. Sequence analysis of the proximal promoter region revealed the presence of a TATA box and putative transcription factor-binding sites, such as those for NF-kappaB and NF-IL6. This region is sufficient to drive high-level reporter gene expression in LPS-stimulated transfected cells.

Genomic organization and promoter analysis of human KCNN3 gene.

KCNN3 is a member of the gene family, KCNN1-4, encoding the small and intermediate conductance calcium-activated potassium channels. Long CAG-repeat alleles of this gene have been found to be over-represented in patients with schizophrenia in a number of population-based association studies, and this gene maps to human chromosome 1q21, a region recently implicated in schizophrenia by linkage. To set the stage for a further functional evaluation of KCNN3, we defined the nature of the genomic locus in the size, structure, and sequence of its introns and exons and the function of potential upstream regulatory regions. We isolated P1-derived artificial chromosome (PAC) clones from a genomic library and identified an overlapping available bacterial artificial chromosome (BAC) clone. Cosmids subcloned from the PAC and BAC clones were then sequenced and merged with the sequence in the public database. The KCNN3 gene spans over 163.1 kb and is composed of eight exons and seven introns. All of the exon-intron junctions conform closely to consensus splice sites. The proximal 2.5 kb of the 5'-flanking sequence was obtained and analyzed for potential transcription factor binding sites. In the proximal 2.5 kb upstream region, potential sites for the Ikaros factor (IK2), homeodomain factor Nkx-2.5/Csx (NKX25), nuclear factor of activated T-cells (NFAT), upstream stimulating factor (USF), c-AMP responsive element binding protein (CREB), POU factor Brn2 (BRN-2), myeloid zinc finger protein (MZF1), vitellogenin binding protein (VBP), HNF3 forkhead homologue 2 (HFH2), and transcription initiation were identified, as well as several potential AP-1 and AP-4 sites. Finally, a 2261-bp fragment of this upstream region was cloned into a promoterless pGL3-luciferase vector, where it produced orientation-dependent expression of the reporter gene in transiently transfected PC12 cells, cells which natively express functional KCNN3 channels, suggesting that this cloned fragment includes competent promoter elements of this gene.

Human RNA Helicase II/Gu Gene: Genomic Organization and Promoter Analysis

Human RNA helicase II/Gu (hRH II/Gu) protein unwinds double-stranded RNA, folds single-stranded RNA, and may play important roles in ribosomal RNA biogenesis, RNA editing, RNA transport, and general transcription. The genomic organization and the regulation of expression of the hRH II/Gu gene were analyzed. The hRH II/Gu gene is over 29 kb in length, and includes 15 exons. All intron–exon junctions contain consensus GT–AG sequences. An 85 bp basic promoter was identified which lacks a functional TATA box. In LNCaP cells, the hRH II/Gu promoter was up to 50 times as active as pGL2-control vector which has an SV 40 early promoter. Mutation of a putative c-Myc/USF binding site in the region between −110 and −85 caused a decrease in the promoter activity by 2- to 3-fold. A negative regulatory region was also found between −324 and −264. Studies of transcriptional regulation of hRH II/Gu under different physiological and pathophysiological conditions may provide clues to its biological function.

Genomic organization and transcriptional analysis of STDEFICIENS in Solanum tuberosum L

The genomic organization of STDEFICIENS (STDEF), the potato orthologous gene to DEFICIENS (DEF) from Antirrhinum majus and APETALA3 (AP3) from Arabidopsis thaliana, has been investigated. Southern-blot analysis on genomic DNA from dihaploid potato lines, using 5′-gene specific probes, revealed polymorphisms that were consistent with the existence in potato of at least two copies of STDEF per haploid genome. This was confirmed by the detection of at least six different STDEF transcripts in the common tetraploid potato S. tuberosum. Genes for two of the STDEF loci, here designated as STDEF-1 and STDEF-2, have been identified as corresponding to the previously described pD13 and pD12 genomic clones, respectively (Garcı́a-Maroto et al., 1993). In addition we have characterised the transcriptional STDEF unit. The main transcription start has been identified around 90 nt upstream of the putative initiation ATG codon, at a CAAATC motif, conserved in AP3. An additional transcription initiation site was detected by 5′-RACE analysis about 300 nt upstream of the main start, which has been confirmed by reverse transcriptase - polymerase chain reaction (RT-PCR) amplification from the longer transcripts. A comparison of the promoter regions for pD12, pD13 and AP3 indicates a similar overall structure, but reveals the existence of a great divergence between pD12 and pD13 in a promoter region that should contain important cis-regulatory elements. This raises the possibility of a differential regulation for the two STDEF genes.

Genomic organization, chromosome location, and expression analysis of mouse β-synuclein, acandidate for involvement in neurodegeneration

The synuclein family of proteins is a group of primarily brain-expressed polypeptides that show a high degree of amino acid conservation. α-Synuclein is the best known of the synuclein family, as it is a major component of the Lewy body, a cytoplasmic inclusion characteristic of Parkinson’s disease as well as a variety of related neurodegenerative disorders. With the discovery that mutations in α-synuclein can cause Parkinson’s disease, a potential role for the other synuclein family members in neurodegenerative disease is being considered. β-Synuclein in particular may deserve special attention, as it is co-expressed with α-synuclein at presynaptic nerve terminals, is subject to phosphorylation by Ca²⁺ calmodulin protein kinase II, appears important for neural plasticity, and forms aggregates in the brains of patients with Parkinson’s disease and a related disorder. To facilitate study of β-synuclein, we have cloned the mouse β-synuclein gene (Sncb) and determined its genomic organization, size, and intron-exon structure. Using an interspecific backcross mapping panel from The Jackson Laboratory, we were then able to localize Sncb to chromosome 13 at the MGD 35.0 cM position. Like the human β-synuclein gene, Sncb appears to consist of six exons separated by five introns. Unlike the human β-synuclein gene, the mouse ortholog possesses a variant GC 5′ splice donor sequence at the exon 4 – intron 4 boundary in a highly conserved splice junction consensus. Northern blot analysis and Western blot analysis both indicate that Sncb is highly expressed in the brain. Knowledge of the genomic organization and expression pattern of Sncb will allow functional studies of its potential role in neurodegeneration to commence in the mouse.

Nucleotide sequence, genome organization and phylogenetic analysis of pineapple mealybug wilt-associated virus-2

The genome of pineapple mealybug wilt-associated closterovirus-2 (PMWaV-2) was cloned from double-stranded RNA isolated from diseased pineapple and its sequence determined. The 3'-terminal 14861 nt of the single-stranded RNA genome contains ten open reading frames (ORFs) which, from 5' to 3', potentially encode a >204 kDa polyprotein containing papain-like protease, methyltransferase and helicase domains (ORF1a), a 65 kDa RNA-dependent RNA polymerase (ORF1b), a 5 kDa hydrophobic protein (ORF2), a 59 kDa heat shock protein 70 homologue (ORF3), a 46 kDa protein (ORF4), a 34 kDa coat protein (ORF5), a 56 kDa diverged coat protein (ORF6), a 20 kDa protein (ORF7), a 22 kDa protein (ORF8) and a 6 kDa protein (ORF9). A 132 nt untranslated region was present at the 3' terminus of the genome. This genome organization is typical of the monopartite closteroviruses, including the putative +1 ribosomal frameshift allowing expression of ORF1b. Phylogenetic analysis revealed that within the family CLOSTEROVIRIDAE: the mealybug-transmitted PMWaV-2 is more closely related to other mealybug-transmitted members than to those which are transmitted by aphids or whiteflies. Within this group, PMWaV-2 shares the greatest sequence identity with grapevine leafroll-associated virus-3, another mealybug-transmitted closterovirus.

Genomic Organization, Physical Mapping, and Expression Analysis of the Human Protein Arginine Methyltransferase 1 Gene

Protein arginine methyltransferases (PRMTs) regulate mRNA processing and maturation by modulating the activity of RNA-binding proteins through methylation. The cDNA for human PRMT1 (HRMT1L2) was recently identified. In this paper, we describe the complete genomic organization of the human PRMT1 gene (GenBank Accession No. AF222689), together with its precise chromosomal localization in relation to other neighboring genes. We have also examined its expression in a total RNA panel of 26 human tissues, the BT-474 breast carcinoma cell line, and 16 breast tumors. PRMT1, which spans 11.2 kb of genomic sequence on chromosome 19q13.3, is located in close proximity to the IRF3 and RRAS genes and is transcribed in the opposite direction. It is formed of 12 coding exons and 11 intervening introns, and shows structural similarity to other PRMT genes. Three PRMT1 isoforms exist as a result of alternative mRNA splicing. Amino acid sequence comparison of the splicing variants indicates that they are all enzymatically active methyl transferases, but with different N-terminal hydrophobic regions. PRMT1 expression was detected in a variety of tissues. We have shown that the relative prevalence of alternatively spliced forms of PRMT1 is different between normal and cancerous breast tissues. Although PRMT1 was not found to be hormonally regulated by steroid hormones in breast cancer cells, our results suggest that two variants of PRMT1 are down regulated in breast cancer.

Human serine racemase: moleular cloning, genomic organization and functional analysis

High levels of d-serine are found in mammalian brain, where it is an endogenous agonist of the strichinine-insensitive site of N-methyl d-aspartate type of glutamate receptors. d-serine is enriched in protoplasmic astrocytes that occur in gray matter areas of the brain and was shown to be synthesized from l-serine . We now report cloning and expression of human serine racemase, an enzyme that catalyses the synthesis of d-serine from l-serine. The enzyme displays a high homology to the murine serine racemase. It contains a pyridoxal 5′-phosphate attachment sequence similar to bacterial biosynthetic threonine dehydratase. Northern-blot analysis show high levels of human serine racemase in areas known to contain large amounts of endogenous d-serine, such as hippocampus and corpus callosum. Robust synthesis of d-serine was detected in cells transfected with human serine racemase, demonstrating the conservation of d-amino acid metabolism in humans. Serine racemase may be a therapeutic target in psychiatric diseases as supplementation of d-serine greatly improves schizophrenia symptoms. We identify the human serine racemase genomic structure and show that the gene encompasses seven exons and localizes to chromosome 17q13.3. Identification of the intron–exon boundaries of the human serine racemase gene will be useful to search for mutations in neuropsychiatric disorders.

Cloning and Characterization of the Gene Encoding the Mouse Peptide Transporter PEPT2

Here we describe the cDNA structure, genomic organization, chromosomal localization, and promoter analysis of the mouse peptide transporter PEPT2. The PEPT2-cDNA is 3987 bp long and encodes a protein of 729 amino acids. The functional properties, analyzed by expression in Xenopus laevis oocytes, showed a typical PEPT2-phenotype with electrogenic, proton-coupled transport, high substrate affinity, and a broad specificity. Immunoblotting of renal brush-border membranes revealed an apparent molecular mass of PEPT2 of 100 kDa. The murine Pept2 gene was cloned from a 129/SvevTACfBr genomic library. It is 34 kb long and comprises 22 exons and 21 introns. By radiation mapping analysis the Pept2 gene was mapped on central mouse chromosome 16. Two putative transcription start sites lying 35 and 235 bp upstream from the translation start were identified. The Pept2 gene possesses a TATA-less promoter. Functional promoter analysis revealed the core promoter to be located between 432 and 286 bp upstream from the translation start.

Genomic organization and promoter analysis of the murine Id3 gene

Overexpression of the helix–loop–helix motif-containing transcription inhibitor Id3 has been shown to repress muscle-specific gene expression. Consistent with its putative negative regulatory role in the myogenic process, Id3 is highly expressed in proliferating myoblasts but down regulated when myoblasts are induced to differentiate. To investigate how Id3 expression may be transcriptionally regulated, we isolated a mouse Id3 genomic DNA fragment and characterized its organization and promoter activity. Comparison of the Id3 gene from human and mouse demonstrated a conserved exon–intron organization in which the first intron interrupts the C-terminal protein coding region and the second intron interrupts the 3′ untranslated region at analogous positions in the two species. Sequence analysis of the 5′-flanking region revealed an unexpected mouse strain-specific genetic polymorphism due to a single base substitution. Deletion analysis revealed that as little as 180 base pairs of the mouse Id3 promoter upstream of the transcription start site is sufficient for a high level of gene expression in proliferating C2C12 myoblasts. In particular, the region between the nucleotide position −180 and −34 appeared to be crucial for maximal reporter gene activity and interacted specifically with C2C12 nuclear proteins. Finally, we showed that, despite the creation of a putative transcription factor-binding site, the genetic polymorphism observed did not affect Id3 promoter activity in proliferating C2C12 cells.

The human Per1 gene: genomic organization and promoter analysis of the first human orthologue of the Drosophila period gene

Per genes encode components of the circadian clocks controlling metabolic and behavioural rhythms. The human Per1 cDNA, RIGUI, was previously isolated and mapped on chromosome 17p12 (Sun, Z.S., Albrecht, U., Zhuchenko, O., Bailey, J., Eichele, G., Lee, C.C., 1997. RIGUI, a putative mammalian orthologue of the Drosophila period gene. Cell 90, 1003–1011). We have now isolated the entire genomic locus containing the human Per1 gene, in a search for genes associated with CpG-rich sequences. The hPer1 gene spans 15 kb of human genomic DNA and is composed of 23 exons, flanked by 5′ and 3′ regulatory regions. Comparison of the hPer1 genomic clone with the dbEST database revealed homologies with putative alternative transcripts. Functional mapping within the 5′ CpG-rich regulatory region enabled us to locate the hPer1 promoter core in a 510 bp-long sequence centred around a TATA box, which supports high levels of hPer1 transcription. A second regulatory region was formally identified in intron 1, which appears to exert a negative role in transcriptional control of hPer1. These regions may be differentially involved in tissue-specificity, and/or circadian regulation, of the human hPer1 gene transcription.