Open AccessRegulated high level expression of a human gamma-globin gene introduced into erythroid cells by an adeno-associated virus vector.
Author(s) -
Christopher E. Walsh,
Johnson M. Liu,
Xianmin Xiao,
Neal S. Young,
Arthur W. Nienhuis,
R. Jude Samulski
Publication year - 1992
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.89.15.7257
Subject(s) - locus control region , adeno associated virus , biology , microbiology and biotechnology , globin , gene , gene cluster , plasmid , locus (genetics) , parvovirus , gene expression , genetic enhancement , virus , recombinant dna , virology , vector (molecular biology) , genetics , promoter
Gene therapy of severe hemoglobinopathies will require high-level expression of a transferred globin gene in erythroid cells. Distant regulatory elements flanking the beta-globin gene cluster, the locus control region, are needed for appropriate expression. We have explored the use of a human parvovirus, the adeno-associated virus (AAV), for globin gene transfer. The human A gamma-globin gene, linked to hypersensitivity site 2 from the locus control region of the beta-globin gene cluster, was subcloned into a plasmid (psub201) containing the AAV inverted terminal repeats. This construct was cotransfected with a helper plasmid containing trans-acting AAV genes into human 293 cells that had been infected with adenovirus. The recombinant AAV vector containing hypersensitivity site 2 stably introduced on average one or two unrearranged proviral copies into human K562 erythroleukemia cells. The transferred globin gene exhibited normal regulation upon hemin induction of erythroid maturation and was expressed at a level equivalent to a native chromosomal A gamma-globin gene.