Initial function analysis of a novel erythroid differentiation related gene EDRF1

Erythroid differentiation depends on the establishment of specific patterns of gene expression. Hypersensitive site 2 (HS2, serving as a major enhance r of globin genes)-binding proteins may be involved in its natural open chr omosomal environment formation. Previously we prepared monoclonal antibodie s against HS2-binding nuclear proteins of terminal differentiated erythroid cells. By utilizing the monoclonal antibodies, we screened lambda -gt11 hu man fetal liver cDNA expression library and obtained one cDNA clone, which was named erythroid differentiation related gene (EDRF1, Genbank accession number AF040247), encompassing an entire open reading frame. We investigate d the expression pattern of EDRF1 by RT-PCR technique. And a clue to the fu nction of EDRF1 has been found from confirmation of high levels of EDRF1 mR NA in differentiated K562 and human fetal liver tissue. To illuminate the f unction of EDRF1 in K562 cells, sense and antisense EDRF1 constructs were p repared and transfected into K562 cells. (X-globin mRNA was down-regulated and EpoR (erythropoietin receptor) mRNA expression was increased in antisen se transfected cells. Cells transfected with sense construct grew more slow ly than control cells suggested by [H-3] thimidine incorporation experiment s. Suppression of K562 proliferation was accompanied by increased spontaneo us hemoglobin synthesis demonstrated by spectrometry. K562 cells transfecte d with sense construct exhibited reduced clongenicity compared with control cells in methycellulose culture. These data provided the evidence that EDR F1 can influence globin expression and hemoglobin synthesis in K562 cells a nd modulated self-renewal in K562 cells.