Molecular mechanism of high hemoglobin F production in Southeast Asian-type hereditary persistence of fetal hemoglobin

Abstract

Hereditary persistence of fetal hemoglobin (HPFH) is associated with a high level of hemoglobin F (HbF) synthesis in adult heterozygotes. In this study, 2 of 6 unrelated HPFH Thai families were found to be Southeast Asian-type HPFH (SEA-HPFH) by analyses of the hematologic data and Southern blot hybridization with polymerase chain reaction-amplified DNA probes. DNA mapping with a probe for a δ-globin fragment showed a 27-kb deletion of DNA that included the β-globin gene and the 3′ deoxyribonuclease I hypersensitive site 1 (3′HS1) sequence downstream. Deletion of the insulator, 3′HS1, and the juxta-position of the HPFH-3 core enhancer downstream to the 3′ breakpoint have been postulated to be the cause of high HbF production in these individuals. To test this hypothesis, we transfected K562 cells with 4 different bacterial artificial chromosome constructs containing the enhanced green fluorescent protein (EGFP) gene at the position of theAγ- globin gene (pEBAC/148β:EGFP). Flow cytometry was used to compare EGFP expression from the pEBAC/148:EGFP construct with the HPFH-3 core enhancer immediately 5′ to the SEA-HPFH breakpoint (pEnH), from the pEBAC/148β:EGFP construct with 8 kb of the breakpoint sequence and the HPFH-3 core enhancer (pSEA-HPFH), and from the construct with 3′HSl followed by the pSEA-HPFH sequence (pSEA-HPFH_3pHS1). The results show that high HbF production in SEA-HPFH occurs from a deletion of the 3′HS1 sequence and the juxtaposition of the HPFH-3 enhancer downstream to the γ-globin gene. ©2006 The Japanese Society of Hematology.