Studies on intracellular lipid droplet-associated proteins : Its regulatory mechanism and clinical application

脂肪分化相关蛋白(AdRP)是一种细胞内脂滴相关蛋白，广泛表达于肝细胞和巨噬细胞中，参与细胞内脂质蓄积。佛波酯(PMA)可刺激小鼠巨噬细胞RAW264.7细胞ADRP的表达。PMA还能刺激该细胞中adrp启动子的活性，其启动子区域位于-2090到-2005bp之间。当突变引入该区域的PU.1/AP-1复合元件时，启动子活性降低。凝胶迁移率移动分析(EMSA)表明，PU1和AP-1与该元件结合。另一方面，PPARγ配体如曲格列酮可刺激小鼠肝细胞NMuLi细胞中ADRP的表达。PPARγ配体的作用也是由PU1/AP-1元件介导的，因为该元件的突变降低了这种作用。在EMSA中，当使用曲格列酮处理的NMuLi细胞的核提取液时，AR-1与该元件形成的复合体增加。蛋白酪氨酸激酶和蛋白激酶C的抑制剂不能抑制曲格列酮诱导的AR-1基因的表达，提示γ配体通过P13蛋白激活AR-1的转录活性。这些结果表明，adrp基因在启动子中通过相同的增强子受到不同的调控。

Adipose differentiation-related protein (ADRP) is an intracellular, lipid droplet-associated protein, being expressed ubiquitously including hepatocytes and macrophages, and is involved in intracellular lipid accumulation. Phorbol ester (PMA) stimulated ADRP expression in mouse macrophage RAW264.7 cells. PMA also stimulated ADRP promoter activity in this cell line, and the responsible promoter region was identified between -2090 and -2005bp. When mutations were introduced into the PU.1/AP-1 composite element in this region, the promoter activity decreased. Using gel mobility shift analyses (EMSA), PU.1 as well as AP-1 were shown to bind conjointly bind to this element. On the other hand, PPARγ ligands such as troglitazone stimulated ADRP expression in mouse hepatocyte NMuLi cells. The effect of PPARγ ligands was also mediated by the PU.1/AP-1 element, since mutations in this element decreased the effect. In EMSA complex formation by AR-1 with this element increased when nuclear extracts from troglitazone-treated NMuLi cells were used. A P13 kinase inhibitor, but not inhibitors of MEK and PKC, inhibited troglitazone-induced expression of the ADRP mRNA., thus indicating that PPARγ ligands stimulated AR-1 transcriptional activity through P13 kinase. All these results indicated that the ADRP gene is regulated differently through the same enhancer in the promoter.