RESUMO
Friedreich ataxia and ataxia with selective vitamin E deficiency (AVED) share very similar clinical phenotypes. We have mapped the AVED locus to proximal 8q with only three large consanguinous Tunisian families, representing to our knowledge the first use of homozygosity mapping for primary linkage analysis. Subsequently, three additional families showed linkage with the same markers. A maximum lod score of 17.9 was obtained at theta = 0 for the haplotype D8S260-D8S510, consisting of the two closest markers. With only 6 families, the AVED locus is therefore mapped precisely as illustrated by the lod-1 confidence interval of 2.4 cM on either side of D8S260-D8S510. Isolation of a yeast artificial chromosome contig > 800 kilobases (kb) showed that D8S260 and D8S510 are less than 400 kb apart.
Assuntos
Cromossomos Humanos Par 8 , Ataxia de Friedreich/genética , Homozigoto , Deficiência de Vitamina E/genética , Adulto , Mapeamento Cromossômico , Cromossomos Artificiais de Levedura , Feminino , Ligação Genética , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , TunísiaRESUMO
Adrenoleukodystrophy (ALD) is a demyelinating disease of the central nervous system that results from a genetic deficiency of ALDP, an ABC protein involved in the transport of very long-chain fatty acids (VLCFAs). The cloning of the ALD gene and the positive effects of allogeneic bone marrow transplantation support the feasibility of a gene therapy approach. We report the retroviral transfer of the ALD cDNA to peripheral blood and bone marrow CD34+ cells from control donors and ALD patients. Prestimulation of these cells with cytokines, followed by infection with the M48-ALD retroviral vector, resulted in 20% transduction efficiency (4-40%) and expression of the vector-encoded ALDP in 20% of CD34+ cells (7.3-50%). Long-term culture (LTC) of transduced CD34+ cells from two ALD patients showed efficient transduction (24-28%) and stable expression (25-32%) of ALDP in derived clonogenic progenitors at 3 weeks of culture. The expression of ALDP in CFU cells derived from 5 and 6 weeks of LTC confirmed the effective transduction of LTC-initiating cells. Expression of ALDP was observed in CD68+ CFU-derived cells, suggesting that monocyte-macrophages, the target bone marrow cells in ALD, were produced from transduced progenitor cells. VL-CFA content was corrected in LTC and CFU-derived cells in proportion to the percentage of transduced cells, indicating that the vector-encoded ALDP was functional. Although not efficient yet to allow a clinical perspective, these results demonstrate the feasibility of ALD gene transfer into CD34+ cells of ALD patients.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Adrenoleucodistrofia/genética , Antígenos CD34 , Células da Medula Óssea/metabolismo , Técnicas de Transferência de Genes , Vetores Genéticos , Proteínas de Membrana/genética , Retroviridae , Células 3T3 , Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP , Animais , Técnicas de Cultura de Células , Fatores Estimuladores de Colônias/farmacologia , DNA Complementar , Ácidos Graxos/metabolismo , Humanos , Camundongos , Fatores de Tempo , TransfecçãoRESUMO
The adrenoleukodystrophy and adrenoleukodystrophy related proteins belong to a new family of half ATP-binding cassette transporters which are localized within the peroxisomal membrane and whose functions are still unknown. They could possibly homo- or heterodimerize resulting in transporters with similar or distinct functions. The expression of adrenoleukodystrophy and adrenoleukodystrophy related genes was studied at the mRNA and protein levels in adult mouse tissues and several human cell lines. We found that adrenoleukodystrophy and adrenoleukodystrophy related genes have strikingly different expression in most mouse tissues and human cell lines analyzed, indicating that adrenoleukodystrophy and adrenoleukodystrophy related proteins do not function as obligatory partners but might rather fulfill similar metabolic functions in different tissues.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Expressão Gênica , Proteínas de Membrana/genética , Proteínas/genética , Subfamília D de Transportador de Cassetes de Ligação de ATP , Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Encéfalo/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Proteínas/metabolismo , RNA Mensageiro , Retina/metabolismo , Testículo/metabolismo , Células Tumorais CultivadasRESUMO
Ataxia with vitamin E deficiency (AVED) is an autosomal recessive disease characterized clinically by neurological symptoms with often striking resemblance to those of Friedreich ataxia. This disorder has been reported previously as familial isolated vitamin E deficiency. We have mapped recently the AVED locus to a 5-cM confidence interval on chromosome 8q by homozygosity mapping in six Mediterranean families. We have now analyzed six new and two previously described families and demonstrate genetic homogeneity despite important clinical variability and wide geographic origins. Analysis of nine new tightly linked microsatellite markers, including four characterized in this study, revealed a predominant but not unique mutation in northern African populations, where this condition is more frequent. Haplotype analysis but also classical recombinations allowed us to refine the AVED position to a 1-cM interval. A YAC contig over this interval was constructed from marker STSs and YAC fingerprint data, in order to facilitate the search of the AVED gene.