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Influence of the polymorphisms of the α-major regulatory element HS-40 on in vitro gene expression

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Autor(es):
D.M. Ribeiro [1] ; T.R. Zaccariotto [2] ; M.N.N. Santos [3] ; F.F. Costa [4] ; M.F. Sonati [5]
Número total de Autores: 5
Afiliação do(s) autor(es):
[1] Universidade Estadual de Campinas. Departamento de Patologia Clínica
[2] Universidade Estadual de Campinas. Departamento de Patologia Clínica
[3] Universidade Estadual de Campinas. Departamento de Patologia Clínica
[4] Universidade Estadual de Campinas. Faculdade de Ciências Médicas. Hemocentro e Departamento de Clínica Médica - Brasil
[5] Universidade Estadual de Campinas. Departamento de Patologia Clínica
Número total de Afiliações: 5
Tipo de documento: Artigo Científico
Fonte: Brazilian Journal of Medical and Biological Research; v. 42, n. 9, p. 783-786, 2009-07-31.
Assunto(s):Polimorfismo genético
Resumo

The α-MRE is the major regulatory element responsible for the expression of human α-like globin genes. It is genetically polymorphic, and six different haplotypes, named A to F, have been identified in some population groups from Europe, Africa and Asia and in native Indians from two Brazilian Indian tribes. Most of the mutations that constitute the α-MRE haplotypes are located in flanking sequences of binding sites for nuclear factors. To our knowledge, there are no experimental studies evaluating whether such variability may influence the α-MRE enhancer activity. We analyzed and compared the expression of luciferase of nine constructs containing different α-MRE elements as enhancers. Genomic DNA samples from controls with A (wild-type α-MRE) and B haplotypes were used to generate C-F haplotypes by site-directed mutagenesis. In addition, three other elements containing only the G→A polymorphism at positions +130, +199, and +209, separately, were also tested. The different α-MRE elements were amplified and cloned into a plasmid containing the luciferase reporter gene and the SV40 promoter and used to transiently transfect K562 cells. A noticeable reduction in luciferase expression was observed with all constructs compared with the A haplotype. The greatest reductions occurred with the F haplotype (+96, C→A) and the isolated polymorphism +209, both located near the SP1 protein-binding sites believed not to be active in vivo. These are the first analyses of α-MRE polymorphisms on gene expression and demonstrate that these single nucleotide polymorphisms, although outside the binding sites for nuclear factors, are able to influence in vitro gene expression. (AU)

Processo FAPESP: 02/13801-7 - Alterações hereditárias das hemoglobinas: genética molecular, aspectos da evolução clínica e produção de animais transgênicos
Beneficiário:Fernando Ferreira Costa
Modalidade de apoio: Auxílio à Pesquisa - Temático
Processo FAPESP: 03/07412-0 - Influencia dos haplotipos do elemento regulatorio do cluster alfa na expressao dos genes da globina alfa humana.
Beneficiário:Daniela Maria Ribeiro
Modalidade de apoio: Bolsas no Brasil - Doutorado