Nemoto Y, Terajima M, Shoji W, Obinata M
Department of Cell Biology, Institute of Development, Aging and Cancer, Tohoku University, Seiryomachi 4-1, Aoba-ku, Sendai 980-77, Japan.
J Biol Chem. 1996 Jun 7;271(23):13542-8. doi: 10.1074/jbc.271.23.13542.
The far upstream region (-1.2-0.9 kilobase pairs) of the mouse glycophorin gene contains the locus control region (LCR)-like region, which acts as an erythroid-specific enhancer dependent on chromosomal integration in murine erythroleukemia (MEL) cells. In the present study, we demonstrated that this region binds six nuclear factors. The binding of GATA-1 to corresponding sites did not show any change before or after induction with dimethyl sulfoxide, but the binding of Spi-1/PU.l and an unidentified factor called glycophorin regulatory element binding factor (GRBF) showed a change during induction. While binding activity of Spi-l/PU.l dropped soon after induction, the GRBF activity increased after induction when expression of the glycophorin gene began. After identification of the consensus binding site of GRBF, we cloned cDNA for that factor by Southwestern method, and it was identified as a previously reported transcription factor, delta, a murine form of YY-l which is a versatile transcription factor. Mutation analysis in the delta/YY-1 binding site within the LCR-like region indicated that delta/YY-1 acts as a regulatory protein in combination with the E-box-binding protein that binds to the neighboring sequence.
小鼠血型糖蛋白基因的远上游区域（-1.2 - 0.9千碱基对）包含类似位点控制区（LCR）的区域，该区域在鼠类红白血病（MEL）细胞中作为依赖于染色体整合的红系特异性增强子发挥作用。在本研究中，我们证明该区域结合六种核因子。在二甲基亚砜诱导前后，GATA-1与相应位点的结合未显示任何变化，但Spi-1/PU.l和一种名为血型糖蛋白调节元件结合因子（GRBF）的未鉴定因子的结合在诱导过程中发生了变化。虽然诱导后不久Spi-1/PU.l的结合活性下降，但当血型糖蛋白基因开始表达时，GRBF活性在诱导后增加。在鉴定出GRBF的共有结合位点后，我们通过蛋白质印迹法克隆了该因子的cDNA，并且它被鉴定为先前报道的转录因子δ，即YY-1的鼠类形式，YY-1是一种多功能转录因子。对类似LCR区域内δ/YY-1结合位点的突变分析表明，δ/YY-1与结合相邻序列的E盒结合蛋白结合，作为一种调节蛋白发挥作用。
