DNA-Protein Interaction of gamma-Globin Gene Promoter by Differentiation Inducers

Kim, Deok In; Park, Joo In; Kim, In Hoo

Abstract

BACKGROUND
The K562 erythroleukemia cell line was used to study the molecular mechanisms regulating lineage commitment of hematopoietic cells. There are numerous similarities between the erythroid or megakaryocytic lineages. In this study, we examined role of the region -269~-240 of gamma-globin gene promoter in fetal hemoglobin expression during either erythroid or megakaryocytic differentiation.
METHODS
K562 cells were cultured and treated with differentiation inducers. Hemoglobin content was scored by benzidine staining, and hemoglobin F was stained by acid elution technique. To determine whether transcription factor binding to the gamma-globin gene promoter is critical to lineage determination, DNA-protein interaction of gamma-globin gene promoter was examined under both uninduced and induced conditions of K562 cells using gel mobility shift assay and southwestern blot analysis.
RESULTS
Phorbol 12-myristate 13-acetate (PMA) induced a megakaryocytic differentiation, but suppressed erythroid differentiation. On the other hand, hydroxyurea (HU), hemin, n-butanol, and sodium butyrate (NaB) induced the expression of erythroid phenotypes. Parallel to hemoglobinization, increase in gamma-globin mRNA was observed in HU- and hemin-treated K562 cells. Gel mobility shift assay and southwestern blot analysis revealed that binding of a erythroid-specific protein (p120) to the region -269~-240 of gamma-globin gene promoter occurred with treatment of erythroid differentiation inducers and did not occur with treatment of PMA.
CONCLUSION
These results suggest that erythroid differentiation inducers may act via DNA- protein interaction at the gamma-globin gene promoter region to induce erythroid differentiation.