Vostrov Alexander A, Taheny Michael J, Quitschke Wolfgang W
Department of Psychiatry and Behavioral Science, State University of New York, Stony Brook, New York 11794-8101, USA.
J Biol Chem. 2002 Jan 11;277(2):1619-27. doi: 10.1074/jbc.M109748200. Epub 2001 Nov 8.
Transcription from the amyloid precursor protein (APP) promoter is largely dependent on a nuclear factor binding site designated as APBbeta. The protein that binds to this site is the multifunctional transcription factor CTCF, which consists of 727 amino acids and contains a domain of 11 zinc finger motifs that is flanked by 267 amino acids on the N-terminal side and 150 amino acids on the C-terminal side. Depleting HeLa cell nuclear extract of endogenous CTCF specifically reduced transcriptional activity from the APP promoter. However, transcriptional activity was restored by replenishing the depleted extract with recombinant CTCF. Deleting 201 amino acids from the C-terminal end of CTCF had no detrimental effect on transcriptional activation, whereas deleting either 248 or 284 amino acids from the N-terminal end abolished transcriptional activation. Competing endogenous CTCF in vivo was accomplished by cotransfecting COS-1 cells with a plasmid overexpressing CTCF constructs and a reporter plasmid containing the APP promoter. Under these conditions, an N-terminal deletion of CTCF reduced expression from the APP promoter, whereas the C-terminal deletion had no effect. These results demonstrate that CTCF activates transcription from the APP promoter and that the activation domain is located on the N-terminal side of the zinc finger domain.
淀粉样前体蛋白（APP）启动子的转录很大程度上依赖于一个被指定为APBβ的核因子结合位点。与该位点结合的蛋白质是多功能转录因子CTCF，它由727个氨基酸组成，包含一个由11个锌指基序组成的结构域，其N端侧翼有267个氨基酸，C端侧翼有150个氨基酸。耗尽HeLa细胞核提取物中的内源性CTCF会特异性降低APP启动子的转录活性。然而，通过用重组CTCF补充耗尽的提取物可恢复转录活性。从CTCF的C末端删除201个氨基酸对转录激活没有不利影响，而从N末端删除248或284个氨基酸则消除了转录激活。在体内竞争内源性CTCF是通过将过表达CTCF构建体的质粒与含有APP启动子的报告质粒共转染COS-1细胞来实现的。在这些条件下，CTCF的N末端缺失会降低APP启动子的表达，而C末端缺失则没有影响。这些结果表明CTCF激活APP启动子的转录，并且激活结构域位于锌指结构域的N端。
