The mechanism of the human β-globin gene switching from an aspect of genomic organization.

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The purpose of the study is to investigate the switching mechanism of the human β-globin gene expression from an aspect of its genomic organization. Five active genes (ε-, Gγ-,Aγ-, δ-and (3-globin genes) exhibit temporal and tissue-specific expression during development. In an attempt to examine the regulation of expression of the β-globin gene family from the basis of DNA structure, we focused on a type of non-B DNA structures, intrinsically bent DNA. Using the circular permutation assay, we mapped the bent DNA sites in the entire region of the β-family globin locus and carried out the detail analyses on the region of bent DNA sites. Based on the previous results, the main object of the study supported by a Grant-in-Aid for Scientific Research (C) was to analyze nucleosomal phases in vitro and in vivo in the DNase I hypersensitive sites (HS)-2 enhancer region of the Locus Control Region (LCR).The HS-2 region was flanked by two DNA bend sites and four nucleosomes were accommodated between the sites. The nucleosomal phases that contained the bend core sequences were determined in vitro and in vivo. On the other hand, the nucleosome in the middle which corresponded to the exact location of HS-2 and included the binding site for the erythroid-specific transcription factor NF-E2 showed several phases in erythroid K562 cells. In contrast, only one phase was adopted in non-erythroid HeLa cells in this region. When the chromatin structure is in a repressive state, the transcriptional efficiency is regulated by controlling the interaction of NF-E2 to its cognate motif on the chromosome before chromatin remodeling. From the results, we hypothesized the function of the periodic bent DNA is a basic factor of key nucleosomes on nucleosome alignment. We currently confirmed the hypothesis by screening and characterizing key nucleosomes using a dinucleosome DNA library.