Distal apolipoprotein C-III regulatory elements F to J act as a general modular enhancer for proximal promoters that contain hormone response elements - Synergism between hepatic nuclear factor-4 molecules bound to the proximal promoter and distal enhancer sites
Distal apolipoprotein C-III regulatory elements F to J act as a general modular enhancer for proximal promoters that contain hormone response elements - Synergism between hepatic nuclear factor-4 molecules bound to the proximal promoter and distal enhancer sites
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DOI：
10.1161/01.atv.17.1.222
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发表时间：
1997-01-01
影响因子：
8.7
通讯作者：
Zannis, V
中科院分区：
文献类型：
--
作者：
Kardassis, D;Tzameli, I;Zannis, V
瞬时转染实验表明，含有调控元件F至J的apoC-III启动子片段可将肝细胞(HepG2)中与其相连的apoA-I近端启动子的强度提高5-13倍。肠道(Caco-2)细胞的激活水平与在HepG2细胞中获得的水平相当，需要一个更大的apoA-I启动子序列，该序列延伸到核苷酸-1500以及肝脏核因子-4(HNF-4)的存在。远端的apoC-III调控元件也可以将异源apoB启动子在HepG2和Caco-2细胞中的强度提高4-8倍。最后，在HNF-4存在的情况下，这些元件将HepG2和Caco-2细胞中与apoA-I的激素反应元件(HRE)AID的两个拷贝相连的最小腺病毒主要晚期启动子的强度提高14.5到18.5倍。启动子/增强子簇的体外突变证实，增强子活性因调节元件I(-736至-714)3‘端的HRE突变而丧失，并因apoC-III增强子的一个或多个调节元件F至J的点突变或缺失而显著降低。增强子的活性还需要apoA-I近端启动子的HRE。ApoC-III增强子还可以恢复因CCAAT/增强子结合蛋白结合位点突变而失活的apoA-I和apoB启动子的活性，表明C/EBP可能不参与启动子/增强子簇的协同激活。这些发现表明，apoC-III启动子的调控元件F到J起到了通用的模块化增强子的作用，可以增强含有HRE的近端启动子的强度。在HepG2细胞中，这种增强作用可以通过HNF-4或与HREs近端和远端HRE结合的Ether核激素受体与SP1或与apoC-III增强子结合的乙醚因子之间的协同作用来解释。Caco-2细胞的最佳活性以及该区域作为肠道增强剂的功能可能需要额外的因素。
Transient transfection assays have shown that the distal apoC-III promoter segments that contain the regulatory elements F to J enhance the strength of the tandemly linked proximal apoA-I promoter 5- to 13-fold in hepatic (HepG2) cells. Activation in intestinal (CaCo-2) cells to levels comparable to those obtained in HepG2 cells requires a larger apoA-I promoter sequence that extends to nucleotide -1500 as well as the presence of hepatic nuclear factor-4 (HNF-4). The distal apoC-III regulatory elements can also enhance 4- to 8-fold the strength of the heterologous apoB promoter in HepG2 and CaCo-2 cells. Finally, these elements in the presence of HNF-4 enhance 14.5- to 18.5-fold the strength of the minimal adenovirus major late promoter linked to two copies of the hormone response element (HRE) AID of apoA-I in both HepG2 and CaCo-2 cells. In vitro mutagenesis of the promoter/enhancer cluster established that the enhancer activity is lost by a mutation in the HRE present in the 3' end of the regulatory element I (-736 to -714) and is reduced significantly by point mutations or deletions in one or more of the regulatory elements F to J of the apoC-III enhancer. The enhancer activity also requires the HREs of the proximal apoA-I promoter. The apoC-III enhancer can also restore the activity of the proximal apoA-I and apoB promoters that have been inactivated by mutations in CCAAT/enhancer binding protein binding sites, indicating that C/EBP may not participate in the synergistic activation of the promoter/enhancer cluster. The findings suggest that the regulatory elements F to J of the apoC-III promoter act as a general modular enhancer that can potentiate the strength of proximal promoters that contain HREs. Such potentiation in the HepG2 cells can be accounted for by synergistic interactions between HNF-4 or Ether nuclear hormone receptors bound to the proximal and distal HREs and SP1 or ether factors bound to the apoC-III enhancer. Additional factors may be required for optimal activity in CaCo-2 cells as well as for the function of this region as an intestinal enhancer.