CLONING AND CHARACTERIZATION OF THE PROMOTER OF THE MURINE LIPOPROTEIN LIPASE-ENCODING GENE - STRUCTURAL AND FUNCTIONAL-ANALYSIS
CLONING AND CHARACTERIZATION OF THE PROMOTER OF THE MURINE LIPOPROTEIN LIPASE-ENCODING GENE - STRUCTURAL AND FUNCTIONAL-ANALYSIS
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DOI：
10.1016/0378-1119(91)90325-6
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发表时间：
1991-11-15
期刊：
影响因子：
3.5
通讯作者：
GIMBLE, JM
中科院分区：
文献类型：
--
作者：
HUA, XX;ENERBACK, S;GIMBLE, JM
脂蛋白脂肪酶（LPL）负责将甘油三酯水解成游离脂肪酸和甘油。当小鼠骨髓基质细胞克隆 BMS2 经历脂肪细胞分化时，会诱导其合成。小鼠基因组 LPL 启动子已被克隆、测序，并通过功能和结构分析进行表征。转录起始点已通过 S1 核酸酶和引物延伸技术绘制。小鼠和人类之间 1.7 kb 的 LPL 5' 侧翼序列的比较显示出 65% 或更高的同一性，并且许多潜在的蛋白质识别基序具有保守性。使用将该区域连接到编码荧光素酶的报告基因的构建体，瞬时转染实验已经记录了该序列在许多细胞系中的启动子功能。基于一组限制性核酸内切酶，紧邻并包括第一个外显子 5'-非翻译区的至少 260 bp 对外源核酸酶消化高度敏感，这与改变的染色质结构一致。根据 ExoIII 足迹和凝胶保留测定，在该区域内的八聚体结合蛋白 1 位点和紧邻翻译起始位点 5' 处检测到蛋白质-DNA 相互作用。
The enzyme lipoprotein lipase (LPL) is responsible for the hydrolysis of triglycerides into free fatty acids and glycerol. Its synthesis is induced as the murine bone marrow stromal cell clone, BMS2, undergoes adipocyte differentiation. The murine genomic LPL promoter has been cloned, sequenced, and characterized by functional and structural assays. The transcriptional start points have been mapped by S1 nuclease and primer extension techniques. Comparison of the 1.7-kb of LPL 5'-flanking sequence between mouse and man reveals 65% identity or greater with conservation of many potential protein-recognition motifs. Using constructs linking this region to the luciferase-encoding reporter gene, transient transfection experiments have documented the promoter function of this sequence in a number of cell lines. Based on a battery of restriction endonucleases, at least 260 bp immediately adjacent to and including the 5'-untranslated region of the first exon are hypersensitive to exogenous nuclease digestion, consistent with an altered chromatin structure. Protein-DNA interactions are detected within this area at the octamer binding protein 1 site and immediately 5' to the translation initiation site based on ExoIII footprinting and gel retention assays.