Kameyama G
First Department of Internal Medicine, Nippon Medical School, Tokyo, Japan.
Nihon Ika Daigaku Zasshi. 1996 Apr;63(2):127-39. doi: 10.1272/jnms1923.63.127.
Locus control region (LCR) is known to occur 5'-upstream of the globin gene clusters in humans and a number of other animals. It comprises four DNase I hypersensitive sites, HS 1-4, and has been considered to play a key role in regulating the globin gene expression in tissue- and developmental stage-specific manners. The occurrence of LCR in the rat genome, however, has not been documented so far. In the present study, the author intended to identify and analyze the rat beta-LCR HS 1 and HS 2, in order to further facilitate studies on the regulatory mechanism involved in globin gene expression. The results obtained in this study are summerized as follows: 1. A DNA region of about 700 bp on the rat genome was amplified by polymerase chain reaction (PCR) using synthetic primers derived from portions of the mouse beta-LCR HS 2. The nucleotide sequence of the PCR product (R 700) shows 67% and 83% homologies with those of the human and mouse HS 2, respectively, indicating that R 700 represents beta-LCR HS 2 of rats. 2. In order to locate beta-LCR HS 2 on the rat genome, a 7 kb DNA fragment (R 7,000) harboring a region between beta-LCR HS 2 and the epsilon 1-globin gene was obtained by PCR. Restriction endonuclease mapping of R 7,000 revealed that the rat beta-LCR HS 2 is located 6.0 kb 5'-upstream relative to the cap site of the epsilon 1-globin gene. 3. The rat beta-LCR HS 1 was then located 4.2 kb 5'-upstream of the epsilon 1-globin gene by Southern blot hybridization of R 7,000 using a human HS 1 probe. Nucleotide sequencing revealed that the rat HS 1 has 83% homology to the mouse HS 1. 4. Comparisons of the structures of the rat beta-LCR HS 1 and HS 2 with those of other animal species indicate that several motifs and consensus sequences for binding of transcription factors, such as NF-E 2/AP-1 and GATA-1, are well conserved during evolutional periods, indicating an indispensable role of LCR in globin gene expression.
基因座控制区（LCR）已知存在于人类和许多其他动物的珠蛋白基因簇的5'上游。它由四个DNA酶I超敏位点HS 1 - 4组成，并被认为在以组织和发育阶段特异性方式调节珠蛋白基因表达中起关键作用。然而，大鼠基因组中LCR的存在迄今尚未见报道。在本研究中，作者旨在鉴定和分析大鼠β-LCR的HS 1和HS 2，以便进一步促进对珠蛋白基因表达调控机制的研究。本研究获得的结果总结如下：1. 使用源自小鼠β-LCR HS 2部分的合成引物，通过聚合酶链反应（PCR）扩增大鼠基因组上约700 bp的DNA区域。PCR产物（R 700）的核苷酸序列与人及小鼠的HS 2分别显示出67%和83%的同源性，表明R 700代表大鼠的β-LCR HS 2。2. 为了在大鼠基因组上定位β-LCR HS 2，通过PCR获得了一个7 kb的DNA片段（R 7,000），其包含β-LCR HS 2和ε1 -珠蛋白基因之间的区域。R 7,000的限制性内切酶图谱显示，大鼠β-LCR HS 位于相对于ε1 -珠蛋白基因帽位点5'上游6.0 kb处。3. 然后通过使用人HS 1探针的R 7,000的Southern印迹杂交，将大鼠β-LCR HS 1定位在ε1 -珠蛋白基因5'上游4.2 kb处。核苷酸测序显示大鼠HS 1与小鼠HS 1具有83%的同源性。4. 大鼠β-LCR HS 1和HS 2与其他动物物种的结构比较表明，转录因子如NF-E 2/AP-1和GATA-1结合的几个基序和共有序列在进化过程中得到了很好的保守，表明LCR在珠蛋白基因表达中具有不可或缺的作用。
