摘要
By electrophoretic mobility shift assay (EMSA), the effect of point mutation C→T at - 64 of human δ-globin gene on its binding proteins has been studied. Two segments of 36 bp from - 83- - 48 bp of the 6 globin gene promoter, named WOG and MOG, were synthesized. WOG includes wild type CAAT-like box (CCAAC), while MOG includes the mutant CAAT-like box (CCAAT, -64 C→T). Results indicate that: ( i ) in erythroid cell lines MEL, K562 and Hemin induced K562, the affinity of MOG with CCAAT binding protein (CBF) and GATA-1
By electrophoretic mobility shift assay (EMSA), the effect of point mutation C→T at ?64 of human 6-globin gene on its binding proteins has been studied. Two segments of 36 bp from ?83-?48 bp of the δ globin gene promoter, named WOG and MOG, were synthesized. WOG includes wild type CAAT-like box (CCAAC), while MOG includes the mutant CAAT-like box (CCAAT, ?64 C → T). Results indicate that: (i) in erythroid cell lines MEL, K562 and Hemin induced K562, the affinity of MOG with CCAAT binding protein (CBF) and GATA-1 was greatly increased; (ii) in Hemin induced K562 cell line, there were another two novel specific DNA binding proteins, named C and D temporarily, besides the above two factors. The former was combined with WOG and MOG, likely indicating its relation with the increased gene expression after induction. The latter was only combined with MOG, which had possible relationship with the point mutation of ?64 C→T; (iii) EMSA also indicates that the suppression mechanisms of the expression of 5 globin gene is different in various periods of human developments. The result evidently supports the hypothesis that the defect CAAT-like box in human 6 globin gene contributes the main reason of its low level expression. The defect c/s-acting element CAAT-like box affects gene expression by its combination with the frans-acting element CBF and GATA-1.
