p300 Coactivates the adipogenic transcription factor CCAAT/enhancer-binding protein α
p300 Coactivates the adipogenic transcription factor CCAAT/enhancer-binding protein α
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DOI：
10.1074/jbc.m100128200
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发表时间：
2001-05-11
影响因子：
4.8
通讯作者：
MacDougald, OA
中科院分区：
文献类型：
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作者：
Erickson, RL;Hemati, N;MacDougald, OA
尽管CCAAT/增强子结合蛋白α（C/EBP α）在前脂肪细胞分化中起着重要作用，但我们对C/EBP α如何与核蛋白相互作用以调节转录的理解有限。基于进化上保守的区域在功能上是重要的并且可能与辅激活因子相互作用的假设，我们比较了来自不同物种（青蛙到人）的C/EBP α的氨基酸序列，并且在反式激活结构域内鉴定了四个高度保守的区域（CR 1-CR 4）。制备了一系列氨基末端截短和内部缺失构建体，产生缺乏单个或多个保守区的C/EBP α形式。为了确定C/EBP α反式激活结构域的哪些区域在其诱导3 T3-L1前脂肪细胞自发分化的能力中是重要的，我们用编码C/EBP α保守区突变体的表达载体感染前脂肪细胞，并观察其诱导分化的能力。我们发现，CR2融合的DNA结合结构域是能够诱导自发分化的其他保守区域独立。然而，CR2对于C/EBP α的脂肪形成作用不是必需的，因为CR 1和CR 3的组合也可以诱导脂肪形成。由于转录辅激活因子p300参与许多转录因子的信号传导到基础转录器，我们研究了C/EBP α和p300之间是否存在功能性相互作用。p300与p42 C/EBP α的共转染导致瘦素启动子活性的协同增加，表明p300作为C/EBP α的转录共激活因子。使用C/EBP α保守区突变体的分析表明，C/EBP α反式激活结构域的多个区域（CR2和CR 3）与p300在功能上相互作用。
Despite the knowledge that CCAAT/enhancer-binding protein alpha (C/EBP alpha) plays an important role in preadipocyte differentiation, our understanding of how C/EBP alpha interacts with nuclear proteins to regulate transcription is limited. Based on the hypothesis that evolutionarily conserved regions are functionally important and likely to interact with coactivators, we compared the amino acid sequence of C/EBP alpha from different species (frog to human) and identified four highly conserved regions (CR1-CR4) within the transactivation domain. A series of amino-terminal truncations and internal deletion constructs were made creating forms of C/EBP alpha which lack single or multiple conserved regions. To determine which regions of the C/EBP alpha transactivation domain are important in its ability to induce spontaneous differentiation of 3T3-L1 preadipocytes, we infected preadipocytes with expression vectors encoding the C/EBP alpha conserved region mutants and observed their ability to induce differentiation. We found that CR2 fused to the DNA binding domain is able to induce spontaneous differentiation independent of the other conserved regions. However, CR2 was not necessary for the adipogenic action of C/EBP alpha because a combination of CR1 and CR3 can also induce adipogenesis. Because the transcriptional coactivator p300 participates in the signaling of many transcription factors to the basal transcriptional apparatus, we examined whether functional interaction exists between C/EBP alpha and p300. Cotransfection of p300 with p42C/EBP alpha results in a synergistic increase in leptin promoter activity, indicating that p300 acts as a transcriptional coactivator of C/EBP alpha. Analyses using C/EBP alpha conserved region mutants suggest that multiple regions (CR2 and CR3) of the C/EBP alpha transactivation domain functionally interact with p300.