the gata1-hs2 enhancer allows persistent and position-independent expression of a β-globin transgenegata1-hs2增强器允许持久和位置无关β-globin转入基因的表达.pdfVIP

The GATA1-HS2 Enhancer Allows Persistent and Position- Independent Expression of a b-globin Transgene 1,2 3 1 1 1 Annarita Miccio , Valentina Poletti , Francesca Tiboni , Claudia Rossi , Antonella Antonelli , Fulvio Mavilio2,3, Giuliana Ferrari1,4* 1 H. San Raffaele-Telethon Institute for Gene Therapy (HSR-TIGET), Istituto Scientifico H. San Raffaele, Milan, Italy, 2 Department of Biomedical Sciences, University of Modena and Reggio Emilia, Modena, Italy, 3 Laboratory of Gene Expression, Istituto Scientifico H. San Raffaele, Milan, Italy, 4 University Vita-Salute San Raffaele, Milan, Italy Abstract Gene therapy of genetic diseases requires persistent and position-independent expression of a therapeutic transgene. Transcriptional enhancers binding chromatin-remodeling and modifying complexes may play a role in shielding transgenes from repressive chromatin effects. We tested the activity of the HS2 enhancer of the GATA1 gene in protecting the expression of a b-globin minigene delivered by a lentiviral vector in hematopoietic stem/progenitor cells. Gene expression from proviruses carrying GATA1-HS2 in both LTRs was persistent and resistant to silencing at most integration sites in the in vivo progeny of human hematopoietic progenitors and murine long-term repopulating stem cells. The GATA1-HS2-modified vector allowed correction of murine b-thalassemia at low copy number without inducing clonal selection of erythroblastic progenitors. Chromatin immunoprecipitation studies showed that GATA1 and the CBP acetyltransferase bind to GATA1-HS2, significantly increasing CBP-specific histone acetylations at the LTRs and b-globin promoter. Recruitment of CBP by the LTRs thus establishes an o