Molecular mechanism of estrogen action to inhibit obesity

雌激素抑制肥胖的分子机制

• 批准号: 08671890
• 负责人: KURACHI Hirohisa
• 金额: $ 1.41万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08671890/
• 关键词: obesity; estrogen; lipoprotein lipase; transcriptional regulation; transcription factor; deletion mutants; mutants; menopause; 脂質代謝; 卵巣摘除マウス; Acyl CoA Synthetase; 上皮成長因子; 分化; Nothern blot

Estrogen is known to regulate adipogenesis in rodents. Estrogen administration in ovariectomized rodents reduces lipoprotein lipase (LPL) gene expression in fat tissues. In this study we studied the mechanisms of transcriptional regulation of the murine LPL gene by estrogen. The 5'-flanking region of LPL gene (-1980 bp) was fused to CAT reporter gene, introduced into COS-1 cells with estrogen receptor expression vector and CAT activities were determined in the presence or the absence of estradiol. Estradiol at 10^<-6> M suppressed pLPL- (-1980) -CAT activity by 20-fold in COS-1 cells. We next determined the estrogen-suppressive element on the LPL promoter by systematically deleting the 5'-flanking region. The most potent region for the estrogen-dependent suppression was located between -1980 and -1570 bp. We narrowed down the region using 50 - 410 bp fragments between -1980 and -1570 bp fused to the minimal promoter-CAT gene. We found that the 50-bp- (-1620/-1570) element was important for estrogen-dependent suppression in LPL gene transcription. When this 50-bp element was deleted from the pLPL (-1980) -CAT construct, estrogen-dependent suppressiveness was decreased to 5-fold. Nuclear proteins from COS-1 cells specifically bound to this 50-bp element were observed by the electrophoretic mobility shift assay. These results collectively suggest that the 50-bp (-1620/-1570) element, which harbors no conventional estrogen-responsive element, may contain a novel cis-acting element which is crucial for estrogen-dependent suppression of the murine LPL gene transcription.

已知雌激素可以调节啮齿动物的脂肪生成。切除卵巢的啮齿动物服用雌激素会降低脂肪组织中脂蛋白脂肪酶（LPL）基因的表达。在这项研究中，我们研究了雌激素对小鼠 LPL 基因转录调控的机制。将LPL基因的5'-侧翼区(-1980 bp)与CAT报告基因融合，用雌激素受体表达载体导入COS-1细胞，并在存在或不存在雌二醇的情况下测定CAT活性。 10^-6M的雌二醇在COS-1细胞中抑制pLPL-(-1980)-CAT活性20倍。接下来，我们通过系统删除 5' 侧翼区域来确定 LPL 启动子上的雌激素抑制元件。雌激素依赖性抑制最有效的区域位于-1980 和-1570 bp 之间。我们使用与最小启动子-CAT 基因融合的 -1980 和 -1570 bp 之间的 50 - 410 bp 片段缩小了区域范围。我们发现 50-bp- (-1620/-1570) 元件对于 LPL 基因转录中雌激素依赖性抑制很重要。当从 pLPL (-1980) -CAT 构建体中删除这个 50 bp 元件时，雌激素依赖性抑制性降低至 5 倍。通过电泳迁移率变动分析观察到与该 50 bp 元件特异性结合的 COS-1 细胞的核蛋白。这些结果共同表明，50 bp (-1620/-1570) 元件不含有传统的雌激素响应元件，但可能含有一种新的顺式作用元件，该元件对于雌激素依赖性抑制小鼠 LPL 基因转录至关重要。