Role of chromatin structure in integration of helper-dependent adenoviral vectors containing the beta-globin locus control region.
We constructed helper-dependent, fiber-chimeric adenoviral vectors that efficiently transduce human hematopoietic stem cells. We found that vectors carrying a 23-kb fragment of the beta-globin locus control region (LCR) flanked by adeno-associated virus inverted terminal repeats (Ad.LCR) preferentially integrated into the chromosomal beta-globin LCR of human erythroid Mo7e cells. We hypothesized that this targeted integration involves beta-globin LCR-specific chromatin structures. Chromatin immunoprecipitation assays of the beta-globin LCR revealed active chromatin within, and immediately downstream of, DNase hypersensitivity region 2 (HS2) in erythroid Mo7e cells, but not in nonerythroid cells. Importantly, most of the Ad.LCR integrations in Mo7e cells were found within this area. We provide further data indicating tethering of incoming Ad.LCR genomes to the chromosomal LCR. We also provide data that suggest a role for active chromatin in AAV Rep78-mediated Ad.LCR integration. Our findings support a new strategy for achieving targeted integration through chromatin tethering of vector DNA.
Sova
Pavel
P
Department of Pathology, University of Washington, Seattle, WA 98195, USA.
Wang
Hongjie
H
Bomsztyk
Karol
K
Stamatoyannopoulos
George
G
Lieber
André
A
D000256:Adenoviridae / Q000235:genetics* / Q000502:physiology; D001665:Binding Sites; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation; D002880:Chromosomes, Human, Pair 11 / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D041905:Erythroid Cells / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics*; D016679:Genome, Viral; D005914:Globins / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006378:Helper Viruses / Q000502:physiology*; D006801:Humans; D019901:Locus Control Region / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D014161:Transduction, Genetic; D014764:Viral Proteins / Q000378:metabolism; D016662:Virus Integration
D002843:Chromatin; D004268:DNA-Binding Proteins; D014764:Viral Proteins; C068734:rep proteins, Adeno-associated virus 2; D049452:Green Fluorescent Proteins; D005914:Globins