Characterization of the human gene encoding the scavenger receptor expressed by endothelial cell and its regulation by a novel transcription factor, endothelial zinc finger protein-2
Characterization of the human gene encoding the scavenger receptor expressed by endothelial cell and its regulation by a novel transcription factor, endothelial zinc finger protein-2
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DOI：
10.1074/jbc.m201854200
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发表时间：
2002-07-05
影响因子：
4.8
通讯作者：
Tsujimoto, M
中科院分区：
文献类型：
--
作者：
Adachi, H;Tsujimoto, M
内皮细胞表达的清道夫受体（SREC）介导内皮细胞选择性摄取修饰的低密度脂蛋白（LDL），如乙酰化LDL和氧化LDL。SREC基因跨越12个内切酶对，包含11个外显子。从外周血白细胞cDNA文库的全长cDNA克隆的SREC的分析显示，至少有五个可变剪接的cDNA存在，其中两个编码可溶性形式的SREC。对SREC基因的转录起始位点进行了定位，DNA序列分析显示在其近端区域有一个Sp1结合位点。5 '侧翼序列的缺失分析表明，碱基对-108和-98之间的序列对启动子活性至关重要。该区域含有一半的反向重复（111）序列和三核苷酸间隔区（IR-3）。利用人脐静脉内皮细胞（HUVEC）核提取物，通过体外DNA酶I足迹分析确定了-268和+17碱基对之间的保护序列。利用单杂交技术从人脐静脉内皮细胞cDNA文库中克隆了一个新的转录因子--内皮锌指蛋白-2（EZF-2），它与SREC启动子活性的5 '侧翼关键区结合。而纯化的重组Sp1.单独在体外DNase I足迹分析中产生类似的保护作用，EZF-2也与5 '侧翼区SREC启动子结合。将SREC启动子与Sp1或EZF-2表达质粒共转染HUVEC，发现EZF-2能增强SREC启动子的活性，而Sp1不能。另一方面，Sp1基序或IR-3基序的突变导致启动子活性降低。这些结果表明，而Sp1是主要的核蛋白结合到启动子的调节区，EZF-2和Sp1负责其调节。
The scavenger receptor expressed by endothelial cell (SREC), mediates the selective uptake of modified low density lipoprotein (LDL), such as acetylated LDL and oxidized LDL, into endothelial cells. The SREC gene spans 12 kilobase pairs and contains 11 exons. Analysis of full-length cDNA clones of SREC from a peripheral blood leukocyte cDNA library revealed that at least five alternatively spliced cDNAs were present, and two of them encoded soluble forms of SREC. The transcription start site of the SREC gene was mapped, and DNA sequence analysis revealed an Sp1 binding site in its proximal region. Deletion analysis of the 5'-flanking sequence revealed that sequence between base pairs -108 and -98 was critical for the promoter activity. This region contained half of an inverted repeat (111) sequence with a triple nucleotide spacer (IR-3). A protected sequence between base pairs -268 and +17 was defined by in vitro DNase I footprinting analysis using human umbilical vein endothelial cell (HUVEC) nuclear extract. A novel transcription factor, endothelial zinc finger protein-2 (EZF-2), that binds to the 5'-flanking critical region of the SREC promoter activity was cloned from a HUVEC cDNA library employing a one-hybrid system. Whereas purified recombinant Sp1. alone produced similar protection in in vitro DNase I footprinting analysis, EZF-2 also bound to the 5'-flanking region SREC promoter. Co-transfection of SREC promoter and Sp1 or EZF-2 expression plasmids in HUVEC revealed that EZF-2 but not Sp1 increased SREC promoter activity. On the other hand, the mutation of either the Sp1 motif or IR-3 motif resulted in a decrease in the promoter activity. These results suggest that whereas Sp1 is the major nuclear protein bound to the regulatory region of the promoter, both EZF-2 and Sp1 are responsible for its regulation.