title: Recombinant erythroid Kruppel-like factor fused to GATA1 up-regulates delta- and gamma-globin expression in erythroid cells.
abstract: The β-hemoglobinopathies sickle cell disease and β-thalassemia are among the most common human genetic disorders worldwide. Hemoglobin A2 (HbA2, α₂δ₂) and fetal hemoglobin (HbF, α₂γ₂) both inhibit the polymerization of hemoglobin S, which results in erythrocyte sickling. Expression of erythroid Kruppel-like factor (EKLF) and GATA1 is critical for transitioning hemoglobin from HbF to hemoglobin A (HbA, α₂β₂) and HbA2. The lower levels of δ-globin expression compared with β-globin expression seen in adulthood are likely due to the absence of an EKLF-binding motif in the δ-globin proximal promoter. In an effort to up-regulate δ-globin to increase HbA2 expression, we created a series of EKLF-GATA1 fusion constructs composed of the transactivation domain of EKLF and the DNA-binding domain of GATA1, and then tested their effects on hemoglobin expression. EKLF-GATA1 fusion proteins activated δ-, γ-, and β-globin promoters in K562 cells, and significantly up-regulated δ- and γ-globin RNA transcript and protein expression in K562 and/or CD34(+) cells. The binding of EKLF-GATA1 fusion proteins at the GATA1 consensus site in the δ-globin promoter was confirmed by chromatin immunoprecipitation assay. Our studies demonstrate that EKLF-GATA1 fusion proteins can enhance δ-globin expression through interaction with the δ-globin promoter, and may represent a new genetic therapeutic approach to β-hemoglobinopathies.
authors.affiliation: Molecular and Clinical Hematology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD; and.
authors.forename: Jianqiong
authors.initials: J
authors.lastname: Zhu
authors.forename: Kyung
authors.initials: K
authors.lastname: Chin
authors.forename: Wulin
authors.initials: W
authors.lastname: Aerbajinai
authors.forename: Cecelia
authors.initials: C
authors.lastname: Trainor
authors.forename: Peter
authors.initials: P
authors.lastname: Gao
authors.forename: Griffin P
authors.initials: GP
authors.lastname: Rodgers
mesh_terms: D018952:Antigens, CD34 / Q000378:metabolism; D001483:Base Sequence; D041905:Erythroid Cells / Q000378:metabolism*; D050982:GATA1 Transcription Factor / Q000378:metabolism*; D017930:Genes, Reporter; D006801:Humans; D020014:K562 Cells; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D015854:Up-Regulation / Q000235:genetics*; D055546:delta-Globins / Q000235:genetics*; D055545:gamma-Globins / Q000235:genetics*
chemical_list: D018952:Antigens, CD34; D050982:GATA1 Transcription Factor; C494688:GATA1 protein, human; D051741:Kruppel-Like Transcription Factors; D011993:Recombinant Fusion Proteins; D055546:delta-Globins; C080774:erythroid Kruppel-like factor; D055545:gamma-Globins