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1.
Bone ; 2018 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-30316000

RESUMO

Analysis of tissue from a 34-years-old male patient from Austrian origin with a history of multiple fractures associated with painful episodes over the carpal, tarsal and at the end of the long bones respectively is presented. Radiographic images and axial 3DCT scans showed widespread defects in trabecular bone architecture and ill-defined cortices over these skeletal sites in the form of discrete cystic-like lesions. Family history indicated two sisters (one half and one full biological sisters) also with a history of fractures. Whole exome sequencing revealed two heterozygous missense mutations in TYROBP (MIM 604142; NM_003332.3) gene encoding for a cell-surface adaptor protein, which is part of a signaling complex triggering activation of immune responses. It is expressed in cells of the ectoderm cell linage such as NK and dendritic cells, macrophages, monocytes, myeloid cells, microglia cells and osteoclasts. The phenotype and genotype of the patient were consistent with the diagnosis of Nasu-Hakola disease (NHD) (OMIM 221770). Investigations at the bone material level of a transiliac bone biopsy sample from the patient using polarized light microscopy and backscatter electron imaging revealed disordered lamellar collagen fibril arrangement and extensively increased matrix mineralization. These findings are the first bone material data in a patient with NHD and point toward an osteoclast defect involvement in this genetic condition.

2.
BMC Med Genet ; 19(1): 125, 2018 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-30041615

RESUMO

BACKGROUND: Cenani-Lenz Syndactyly (CLS) syndrome is a rare autosomal recessive disorder characterized by syndactyly and oligodactyly of fingers and toes, disorganization and fusion of metacarpals, metatarsals and phalanges, radioulnar synostosis and mesomelic shortness of the limbs, with lower limbs usually being much less affected than upper limbs. CASE PRESENTATION: we report here two patients, born to consanguineous Sri Lankan parents, present with bilateral postaxial oligodactyly limited to upper limbs. While the proband has no noticeable facial dysmorphism, renal impairments or cognitive impairments, his affected sister displays a few mild facial dysmorphic features. Whole exome sequencing of the proband showed a novel deleterious homozygous mutation (c.1348A > G) in the LRP4 gene, resulting in an Ile450-to-Val (I450V) substitution. CONCLUSION: This recessive mutation in LRP4 confirmed the diagnosis of CLS syndrome in two patients present with isolated hand syndactyly. This is the first reported case of CLS syndrome in a family of Sri Lankan origin.

3.
Eur J Med Genet ; 61(10): 585-595, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29605658

RESUMO

Mutations in CPLANE1 (previously known as C5orf42) cause Oral-Facial-Digital Syndrome type VI (OFD6) as well as milder Joubert syndrome (JS) phenotypes. Seven new cases from five unrelated families diagnosed with pure OFD6 were systematically examined. Based on the clinical manifestations of these patients and those described in the literature, we revised the diagnostic features of OFD6 and include the seven most common characteristics: 1) molar tooth sign, 2) tongue hamartoma and/or lobulated tongue, 3) additional frenula, 4) mesoaxial polydactyly of hands, 5) preaxial polydactyly of feet, 6) syndactyly and/or bifid toe, and 7) hypothalamic hamartoma. By whole or targeted exome sequencing, we identified seven novel germline recessive mutations in CPLANE1, including missense, nonsense, frameshift and canonical splice site variants, all causing OFD6 in these patients. Since CPLANE1 is also mutated in JS patients, we examined whether a genotype-phenotype correlation could be established. We gathered and compared 46 biallelic CPLANE1 mutations reported in 32 JS and 26 OFD6 patients. Since no clear correlation between paired genotypes and clinical outcomes could be determined, we concluded that patient's genetic background and gene modifiers may modify the penetrance and expressivity of CPLANE1 causal alleles. To conclude, our study provides a comprehensive view of the phenotypic range, the genetic basis and genotype-phenotype association in OFD6 and JS. The updated phenotype scoring system together with the identification of new CPLANE1 mutations will help clinicians and geneticists reach a more accurate diagnosis for JS-related disorders.

4.
Proc Biol Sci ; 284(1863)2017 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-28931746

RESUMO

Studies of the voltage-gated sodium (Nav) channels of extant gnathostomes have made it possible to deduce that ancestral gnathostomes possessed four voltage-gated sodium channel genes derived from a single ancestral chordate gene following two rounds of genome duplication early in vertebrates. We investigated the Nav gene family in two species of lampreys (the Japanese lamprey Lethenteron japonicum and sea lamprey Petromyzon marinus) (jawless vertebrates-agnatha) and compared them with those of basal vertebrates to better understand the origin of Nav genes in vertebrates. We noted six Nav genes in both lamprey species, but orthology with gnathostome (jawed vertebrate) channels was inconclusive. Surprisingly, the Nav2 gene, ubiquitously found in invertebrates and believed to have been lost in vertebrates, is present in lampreys, elephant shark (Callorhinchus milii) and coelacanth (Latimeria chalumnae). Despite repeated duplication of the Nav1 family in vertebrates, Nav2 is only in single copy in those vertebrates in which it is retained, and was independently lost in ray-finned fishes and tetrapods. Of the other five Nav channel genes, most were expressed in brain, one in brain and heart, and one exclusively in skeletal muscle. Invertebrates do not express Nav channel genes in muscle. Thus, early in the vertebrate lineage Nav channels began to diversify and different genes began to express in heart and muscle.


Assuntos
Evolução Molecular , Proteínas de Peixes/genética , Duplicação Gênica , Lampreias/genética , Canais de Sódio Disparados por Voltagem/genética , Animais , Filogenia
5.
Am J Hum Genet ; 101(3): 391-403, 2017 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-28886341

RESUMO

In five separate families, we identified nine individuals affected by a previously unidentified syndrome characterized by growth retardation, spine malformation, facial dysmorphisms, and developmental delays. Using homozygosity mapping, array CGH, and exome sequencing, we uncovered bi-allelic loss-of-function CDK10 mutations segregating with this disease. CDK10 is a protein kinase that partners with cyclin M to phosphorylate substrates such as ETS2 and PKN2 in order to modulate cellular growth. To validate and model the pathogenicity of these CDK10 germline mutations, we generated conditional-knockout mice. Homozygous Cdk10-knockout mice died postnatally with severe growth retardation, skeletal defects, and kidney and lung abnormalities, symptoms that partly resemble the disease's effect in humans. Fibroblasts derived from affected individuals and Cdk10-knockout mouse embryonic fibroblasts (MEFs) proliferated normally; however, Cdk10-knockout MEFs developed longer cilia. Comparative transcriptomic analysis of mutant and wild-type mouse organs revealed lipid metabolic changes consistent with growth impairment and altered ciliogenesis in the absence of CDK10. Our results document the CDK10 loss-of-function phenotype and point to a function for CDK10 in transducing signals received at the primary cilia to sustain embryonic and postnatal development.


Assuntos
Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/fisiologia , Deficiências do Desenvolvimento/genética , Transtornos do Crescimento/genética , Mutação , Coluna Vertebral/anormalidades , Coluna Vertebral/patologia , Animais , Ciclo Celular , Proliferação de Células , Células Cultivadas , Criança , Pré-Escolar , Cílios/metabolismo , Cílios/patologia , Deficiências do Desenvolvimento/patologia , Embrião de Mamíferos/metabolismo , Embrião de Mamíferos/patologia , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Transtornos do Crescimento/patologia , Humanos , Lactente , Masculino , Camundongos , Camundongos Knockout , Linhagem , Fosforilação , Transdução de Sinais , Coluna Vertebral/metabolismo
6.
Proc Natl Acad Sci U S A ; 114(34): 9146-9151, 2017 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-28784804

RESUMO

ParaHox genes (Gsx, Pdx, and Cdx) are an ancient family of developmental genes closely related to the Hox genes. They play critical roles in the patterning of brain and gut. The basal chordate, amphioxus, contains a single ParaHox cluster comprising one member of each family, whereas nonteleost jawed vertebrates contain four ParaHox genomic loci with six or seven ParaHox genes. Teleosts, which have experienced an additional whole-genome duplication, contain six ParaHox genomic loci with six ParaHox genes. Jawless vertebrates, represented by lampreys and hagfish, are the most ancient group of vertebrates and are crucial for understanding the origin and evolution of vertebrate gene families. We have previously shown that lampreys contain six Hox gene loci. Here we report that lampreys contain only two ParaHox gene clusters (designated as α- and ß-clusters) bearing five ParaHox genes (Gsxα, Pdxα, Cdxα, Gsxß, and Cdxß). The order and orientation of the three genes in the α-cluster are identical to that of the single cluster in amphioxus. However, the orientation of Gsxß in the ß-cluster is inverted. Interestingly, Gsxß is expressed in the eye, unlike its homologs in jawed vertebrates, which are expressed mainly in the brain. The lamprey Pdxα is expressed in the pancreas similar to jawed vertebrate Pdx genes, indicating that the pancreatic expression of Pdx was acquired before the divergence of jawless and jawed vertebrate lineages. It is likely that the lamprey Pdxα plays a crucial role in pancreas specification and insulin production similar to the Pdx of jawed vertebrates.


Assuntos
Genes Homeobox/genética , Lampreias/genética , Família Multigênica , Vertebrados/genética , Sequência de Aminoácidos , Animais , Evolução Molecular , Proteínas de Peixes/genética , Perfilação da Expressão Gênica/métodos , Proteínas de Homeodomínio/classificação , Proteínas de Homeodomínio/genética , Filogenia , Homologia de Sequência de Aminoácidos , Vertebrados/classificação
7.
Am J Hum Genet ; 100(4): 659-665, 2017 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-28318499

RESUMO

Arthrogryposis multiplex congenita (AMC) is a developmental condition characterized by multiple joint contractures resulting from reduced or absent fetal movements. Through genetic mapping of disease loci and whole-exome sequencing in four unrelated multiplex families presenting with severe AMC, we identified biallelic loss-of-function mutations in LGI4 (leucine-rich glioma-inactivated 4). LGI4 is a ligand secreted by Schwann cells that regulates peripheral nerve myelination via its cognate receptor ADAM22 expressed by neurons. Immunolabeling experiments and transmission electron microscopy of the sciatic nerve from one of the affected individuals revealed a lack of myelin. Functional tests using affected individual-derived iPSCs showed that these germline mutations caused aberrant splicing of the endogenous LGI4 transcript and in a cell-based assay impaired the secretion of truncated LGI4 protein. This is consistent with previous studies reporting arthrogryposis in Lgi4-deficient mice due to peripheral hypomyelination. This study adds to the recent reports implicating defective axoglial function as a key cause of AMC.


Assuntos
Artrogripose/genética , Proteínas da Matriz Extracelular/genética , Mutação , Células de Schwann/metabolismo , Artrogripose/diagnóstico , Artrogripose/patologia , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Bainha de Mielina/metabolismo , Linhagem
8.
Cilia ; 5: 8, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27069622

RESUMO

BACKGROUND: Endocrine-cerebro-osteodysplasia (ECO) syndrome [MIM:612651] caused by a recessive mutation (p.R272Q) in Intestinal cell kinase (ICK) shows significant clinical overlap with ciliary disorders. Similarities are strongest between ECO syndrome, the Majewski and Mohr-Majewski short-rib thoracic dysplasia (SRTD) with polydactyly syndromes, and hydrolethalus syndrome. In this study, we present a novel homozygous ICK mutation in a fetus with ECO syndrome and compare the effect of this mutation with the previously reported ICK variant on ciliogenesis and cilium morphology. RESULTS: Through homozygosity mapping and whole-exome sequencing, we identified a second variant (c.358G > T; p.G120C) in ICK in a Turkish fetus presenting with ECO syndrome. In vitro studies of wild-type and mutant mRFP-ICK (p.G120C and p.R272Q) revealed that, in contrast to the wild-type protein that localizes along the ciliary axoneme and/or is present in the ciliary base, mutant proteins rather enrich in the ciliary tip. In addition, immunocytochemistry revealed a decreased number of cilia in ICK p.R272Q-affected cells. CONCLUSIONS: Through identification of a novel ICK mutation, we confirm that disruption of ICK causes ECO syndrome, which clinically overlaps with the spectrum of ciliopathies. Expression of ICK-mutated proteins result in an abnormal ciliary localization compared to wild-type protein. Primary fibroblasts derived from an individual with ECO syndrome display ciliogenesis defects. In aggregate, our findings are consistent with recent reports that show that ICK regulates ciliary biology in vitro and in mice, confirming that ECO syndrome is a severe ciliopathy.

9.
Arch Iran Med ; 19(2): 87-91, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26838077

RESUMO

BACKGROUND: TWINKLE (c10orf2) gene is responsible for autosomal dominant progressive external ophthalmoplegia (PEO). In rare cases, additional features such as muscle weakness, peripheral neuropathy, ataxia, cardiomyopathy, dysphagia, dysphonia, cataracts, depression, dementia, parkinsonism, and hearing loss have been reported in association with heterozygous mutations of the TWINKLE gene. METHODS: We have studied a large Iranian family with myopathy, dysphonia, dysphagia, and behavior change in addition to PEO in affected members. RESULTS: We identified a missense mutation c.1121G > A in the c10orf2 gene in all affected members. Early death is a novel feature seen in affected members of this family that has not been reported to date. CONCLUSION: The association of PEO, myopathy, dysphonia, dysphagia, behavior change and early death has not been previously reported in the literature or other patients with this mutation.


Assuntos
DNA Helicases/genética , Transtornos de Deglutição/genética , Disfonia/genética , Transtornos Mentais/genética , Proteínas Mitocondriais/genética , Debilidade Muscular/genética , Oftalmoplegia Externa Progressiva Crônica/genética , Adulto , Análise Mutacional de DNA , Transtornos de Deglutição/complicações , Disfonia/complicações , Exoma , Feminino , Técnicas de Genotipagem , Humanos , Irã (Geográfico) , Masculino , Transtornos Mentais/complicações , Pessoa de Meia-Idade , Debilidade Muscular/complicações , Mutação de Sentido Incorreto , Oftalmoplegia Externa Progressiva Crônica/complicações , Linhagem , Adulto Jovem
10.
Mol Biol Evol ; 33(2): 311-5, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26545918

RESUMO

The brain, comprising billions of neurons and intricate neural networks, is arguably the most complex organ in vertebrates. The diversity of individual neurons is fundamental to the neuronal network complexity and the overall function of the vertebrate brain. In jawed vertebrates, clustered protocadherins provide the molecular basis for this neuronal diversity, through stochastic and combinatorial expression of their various isoforms in individual neurons. Based on analyses of transcriptomes from the Japanese lamprey brain and sea lamprey embryos, genome assemblies of the two lampreys, and brain expressed sequence tags of the inshore hagfish, we show that extant jawless vertebrates (cyclostomes) lack the clustered protocadherins. Our findings indicate that the clustered protocadherins originated from a nonclustered protocadherin in the jawed vertebrate ancestor, after the two rounds of whole-genome duplication. In the absence of clustered protocadherins, cyclostomes might have evolved novel molecules or mechanisms for generating neuronal diversity which remains to be discovered.


Assuntos
Caderinas/genética , Lampreias/anatomia & histologia , Lampreias/genética , Família Multigênica , Animais , Caderinas/química , Ordem dos Genes , Genoma , Humanos , Arcada Osseodentária , Vertebrados
11.
Nature ; 505(7482): 174-9, 2014 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-24402279

RESUMO

The emergence of jawed vertebrates (gnathostomes) from jawless vertebrates was accompanied by major morphological and physiological innovations, such as hinged jaws, paired fins and immunoglobulin-based adaptive immunity. Gnathostomes subsequently diverged into two groups, the cartilaginous fishes and the bony vertebrates. Here we report the whole-genome analysis of a cartilaginous fish, the elephant shark (Callorhinchus milii). We find that the C. milii genome is the slowest evolving of all known vertebrates, including the 'living fossil' coelacanth, and features extensive synteny conservation with tetrapod genomes, making it a good model for comparative analyses of gnathostome genomes. Our functional studies suggest that the lack of genes encoding secreted calcium-binding phosphoproteins in cartilaginous fishes explains the absence of bone in their endoskeleton. Furthermore, the adaptive immune system of cartilaginous fishes is unusual: it lacks the canonical CD4 co-receptor and most transcription factors, cytokines and cytokine receptors related to the CD4 lineage, despite the presence of polymorphic major histocompatibility complex class II molecules. It thus presents a new model for understanding the origin of adaptive immunity.


Assuntos
Evolução Molecular , Genoma/genética , Tubarões/genética , Animais , Cálcio/metabolismo , Linhagem da Célula/imunologia , Proteínas de Peixes/classificação , Proteínas de Peixes/genética , Deleção de Genes , Genômica , Imunidade Celular/genética , Anotação de Sequência Molecular , Dados de Sequência Molecular , Osteogênese/genética , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Filogenia , Estrutura Terciária de Proteína/genética , Tubarões/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia , Fatores de Tempo , Vertebrados/classificação , Vertebrados/genética , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento
12.
Proc Natl Acad Sci U S A ; 110(40): 16044-9, 2013 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-24043829

RESUMO

Cyclostomes, comprising jawless vertebrates such as lampreys and hagfishes, are the sister group of living jawed vertebrates (gnathostomes) and hence an important group for understanding the origin and diversity of vertebrates. In vertebrates and other metazoans, Hox genes determine cell fate along the anteroposterior axis of embryos and are implicated in driving morphological diversity. Invertebrates contain a single Hox cluster (either intact or fragmented), whereas elephant shark, coelacanth, and tetrapods contain four Hox clusters owing to two rounds of whole-genome duplication ("1R" and "2R") during early vertebrate evolution. By contrast, most teleost fishes contain up to eight Hox clusters because of an additional "teleost-specific" genome duplication event. By sequencing bacterial artificial chromosome (BAC) clones and the whole genome, here we provide evidence for at least six Hox clusters in the Japanese lamprey (Lethenteron japonicum). This suggests that the lamprey lineage has experienced an additional genome duplication after 1R and 2R. The relative age of lamprey and human paralogs supports this hypothesis. Compared with gnathostome Hox clusters, lamprey Hox clusters are unusually large. Several conserved noncoding elements (CNEs) were predicted in the Hox clusters of lamprey, elephant shark, and human. Transgenic zebrafish assay indicated the potential of CNEs to function as enhancers. Interestingly, CNEs in individual lamprey Hox clusters are frequently conserved in multiple Hox clusters in elephant shark and human, implying a many-to-many orthology relationship between lamprey and gnathostome Hox clusters. Such a relationship suggests that the first two rounds of genome duplication may have occurred independently in the lamprey and gnathostome lineages.


Assuntos
Evolução Molecular , Genes Homeobox/genética , Lampreias/genética , Família Multigênica/genética , Animais , Sequência de Bases , Cromossomos Artificiais Bacterianos/genética , Sequência Conservada/genética , Japão , Funções Verossimilhança , Modelos Genéticos , Dados de Sequência Molecular , Sequências Reguladoras de Ácido Nucleico/genética , Alinhamento de Sequência , Análise de Sequência de DNA
13.
PLoS Genet ; 8(7): e1002798, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22807687

RESUMO

Heterogametic sex chromosomes have evolved independently in various lineages of vertebrates. Such sex chromosome pairs often contain nonrecombining regions, with one of the chromosomes harboring a master sex-determining (SD) gene. It is hypothesized that these sex chromosomes evolved from a pair of autosomes that diverged after acquiring the SD gene. By linkage and association mapping of the SD locus in fugu (Takifugu rubripes), we show that a SNP (C/G) in the anti-Müllerian hormone receptor type II (Amhr2) gene is the only polymorphism associated with phenotypic sex. This SNP changes an amino acid (His/Asp384) in the kinase domain. While females are homozygous (His/His384), males are heterozygous. Sex in fugu is most likely determined by a combination of the two alleles of Amhr2. Consistent with this model, the medaka hotei mutant carrying a substitution in the kinase domain of Amhr2 causes a female phenotype. The association of the Amhr2 SNP with phenotypic sex is conserved in two other species of Takifugu but not in Tetraodon. The fugu SD locus shows no sign of recombination suppression between X and Y chromosomes. Thus, fugu sex chromosomes represent an unusual example of proto-sex chromosomes. Such undifferentiated X-Y chromosomes may be more common in vertebrates than previously thought.


Assuntos
Substituição de Aminoácidos/genética , Receptores de Peptídeos/genética , Receptores de Fatores de Crescimento Transformadores beta/genética , Processos de Determinação Sexual/genética , Takifugu , Animais , Evolução Biológica , Feminino , Estudos de Associação Genética , Ligação Genética , Heterozigoto , Homozigoto , Masculino , Mutação de Sentido Incorreto/genética , Receptores de Peptídeos/fisiologia , Receptores de Fatores de Crescimento Transformadores beta/fisiologia , Cromossomos Sexuais/genética , Takifugu/genética , Takifugu/fisiologia
14.
Genome Biol Evol ; 3: 424-42, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21551351

RESUMO

The compact genome of fugu (Takifugu rubripes) has been used widely as a reference genome for understanding the evolution of vertebrate genomes. However, the fragmented nature of the fugu genome assembly has restricted its use for comparisons of genome architecture in vertebrates. To extend the contiguity of the assembly to the chromosomal level, we have generated a comprehensive genetic map of fugu and anchored the scaffolds of the assembly to the 22 chromosomes of fugu. The map consists of 1,220 microsatellite markers that provide anchor points to 697 scaffolds covering 86% of the genome assembly (http://www.fugu-sg.org/). The integrated genome map revealed a higher recombination rate in fugu compared with other vertebrates and a wide variation in the recombination rate between sexes and across chromosomes of fugu. We used the extended assembly to explore recent rearrangement events in the lineages of fugu, Tetraodon, and medaka and compared them with rearrangements in three mammalian (human, mouse, and opossum) lineages. Between the two pufferfishes, fugu has experienced fewer chromosomal rearrangements than Tetraodon. The gene order is more highly conserved in the three teleosts than in mammals largely due to a lower rate of interchromosomal rearrangements in the teleosts. These results provide new insights into the distinct patterns of genome evolution between teleosts and mammals. The consolidated genome map and the genetic map of fugu are valuable resources for comparative genomics of vertebrates and for elucidating the genetic basis of the phenotypic diversity of ~25 species of Takifugu that evolved within the last 5 My.


Assuntos
Mapeamento Cromossômico , Evolução Molecular , Genoma , Mamíferos/genética , Takifugu/genética , Animais , Cromossomos , Sequência Conservada/genética , Ordem dos Genes , Rearranjo Gênico , Humanos , Hibridização in Situ Fluorescente , Camundongos , Repetições de Microssatélites , Gambás/genética , Oryzias/genética , Filogenia , Análise de Sequência de DNA , Tetraodontiformes/genética
15.
Mar Genomics ; 3(2): 63-70, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21798198

RESUMO

Animals respond to hypoxia by increasing synthesis of the glycoprotein hormone erythropoietin (Epo) which in turn stimulates the production of red blood cells. The gene encoding Epo has been recently cloned in teleost fishes such as the pufferfish Takifugu rubripes (fugu) and zebrafish (Danio rerio). It has been shown that the transcription levels of Epo in teleost fishes increase in response to anemia or hypoxia in a manner similar to its human ortholog. However, the cis-regulatory element(s) mediating the hypoxia response of Epo gene in fishes has not been identified. In the present study, using the human hepatoma cell line (Hep3B), we have identified and characterized a hypoxia response element (HRE) in the fugu Epo locus. The sequence of the fugu HRE (ACGTGCTG) is identical to that of the HRE in the human EPO locus. However, unlike the HRE in the mammalian Epo locus, which is located in the 3' region of the gene, the fugu HRE is located in the 5' flanking region and on the opposite strand of DNA. This HRE is conserved in other teleosts such as Tetraodon and zebrafish in a similar location. A 365-bp fragment containing the fugu HRE was able to drive GFP expression in the liver of transgenic zebrafish. However, we could not ascertain if the expression of transgene is induced by hypoxia in vivo due to the low and variable levels of GFP expression in transgenic zebrafish. Our investigations also revealed that the Epo locus has experienced extensive rearrangements during vertebrate evolution.


Assuntos
Eritropoetina/metabolismo , Hipóxia/metabolismo , Elementos Reguladores de Transcrição/genética , Takifugu/genética , Takifugu/metabolismo , Animais , Sequência de Bases , Linhagem Celular Tumoral , Sequência Conservada/genética , Primers do DNA/genética , Eritropoetina/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Hipóxia/genética , Luciferases , Dados de Sequência Molecular , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de Sequência , Análise de Sequência de DNA , Peixe-Zebra , beta-Galactosidase
16.
Blood ; 104(5): 1498-503, 2004 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-15142879

RESUMO

In this paper we report the cloning and characterization of the erythropoietin (Epo) gene from the pufferfish, Fugu rubripes. This is the first nonmammalian Epo gene to be cloned. The Fugu Epo comprises 5 exons and 4 introns similar to the human EPO, and encodes a 185-amino acid protein that is 32% to 34% identical to Epo from various mammals. The synteny of genes at the Epo locus is conserved between the Fugu and humans. Unlike in mammals in which adult kidney is the primary Epo-producing organ, the heart is the main Epo-producing organ in adult Fugu. In addition to the heart, Fugu Epo is also expressed in the liver and brain similar to the human EPO. Interestingly, the transcripts in the Fugu brain are generated from a distal promoter and include an alternatively spliced first coding exon. No such brain-specific alternative splicing of Epo has been reported in mammals so far. Transient transfection studies in a fish hepatoma cell line (PLHC-1) and a human hepatoma cell line (HepG2) suggest that although the Fugu Epo promoter many not be hypoxia inducible, the gene may be regulated by hypoxia.


Assuntos
Eritropoetina/genética , Coração/fisiologia , Takifugu/genética , Animais , Clonagem Molecular , Éxons , Expressão Gênica , Humanos , Íntrons , Mamíferos , Dados de Sequência Molecular , Regiões Promotoras Genéticas/genética
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