title: Adeno-associated virus 2-mediated transduction and erythroid cell-specific expression of a human beta-globin gene.
abstract: Recombinant adeno-associated virus 2 (AAV) virions were constructed that contained the genomic copy of a normal human beta-globin gene marked with a 4-bp Clal linker, and the herpesvirus thymidine kinase (TK) promoter-driven bacterial gene for resistance to neomycin (v beta m-globin), as well as those containing the DNase l-hypersensitive site 2 (HS-2) from the locus control region (LCR) of the human beta-globin gene cluster (vHS2-beta m-globin). These recombinant virions were used to infect a human erythroleukemia cell line which normally does not express the beta-globin gene (K562), or a human nasopharyngeal carcinoma cell line (KB). Cell populations resistant to G418, a neomycin analogue, were obtained following infections with the recombinant virions, indicating high-efficiency transduction of the chimeric gene as well as functional activity of the transduced neo gene in both cell types. Southern blot analysis using a human beta-globin DNA probe substantiated stable integration of the exogenous beta-globin allele in these cells. There was no expression of the transduced beta-globin gene in K562 or KB cells infected with the v beta m-globin virus. High-level expression of the transduced beta-globin gene occurred only in the vHS2-beta m-globin virus-infected K562 cells, but not in KB cells, as determined by Northern blot as well as RNase protection analyses. Expression of the human beta-globin protein could also be detected in approximately 10-20% of the vHS2-beta m-globin virus-infected K562 cells. These studies suggest that the AAV-based vector system may prove useful for high-efficiency globin gene transfer in human hematopoietic cells.
authors.affiliation: Department of Medicine, Indiana University School of Medicine, Indianapolis 46202-5120, USA.
authors.forename: S Z
authors.initials: SZ
authors.lastname: Zhou
authors.forename: Q
authors.initials: Q
authors.lastname: Li
authors.forename: G
authors.initials: G
authors.lastname: Stamatoyannopoulos
authors.forename: A
authors.initials: A
authors.lastname: Srivastava
mesh_terms: D000229:Dependovirus / Q000235:genetics*; D005914:Globins / Q000235:genetics*; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000235:genetics* / Q000473:pathology; D011995:Recombination, Genetic; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured
chemical_list: D005914:Globins