Yin L, Zhang Y, Charron T, Hillgartner F B
Department of Biochemistry, School of Medicine, P.O. Box 9142, West Virginia University, Morgantown, WV 26506-9142, USA.
Biochim Biophys Acta. 2000 Dec 15;1517(1):91-9. doi: 10.1016/s0167-4781(00)00267-0.
High-carbohydrate feeding and triiodothyronine (T3) increase the abundance of acetyl-CoA carboxylase-alpha (ACC alpha) mRNA in avian hepatocytes, whereas starvation, glucagon, and medium-chain fatty acids decrease the abundance of ACC alpha mRNA. These changes in ACC alpha mRNA levels are mediated by alterations in the rate of transcription of the ACC alpha gene. In liver, ACC alpha transcription is initiated from two promoters, promoter 1 and promoter 2, resulting in transcripts that contain heterogeneity in their 5'-untranslated regions. Here, we investigated the role of promoter 1 and promoter 2 in mediating nutrient- and hormone-induced changes in ACC alpha mRNA abundance by measuring the level of transcripts expressed from promoter 1 and promoter 2 using a ribonuclease protection assay. The results indicated that both promoter 1 and promoter 2 were regulated by starvation/refeeding in livers of intact chicks and by T3, glucagon, and medium-chain fatty acids in chick embryo hepatocyte cultures and that alterations in the activity of promoter 2 accounted for a greater proportion of the changes in total ACC alpha mRNA abundance caused by nutrient and hormone treatment. Five DNase-hypersensitive sites were also identified between -500 and +1 bp relative to the transcription start site of promoter 2 in livers of intact chicks and in chick embryo hepatocyte cultures. In transient transfection analyses, this region of DNase hypersensitivity conferred regulation of transcription by T3, glucagon, and medium-chain fatty acids in chick embryo hepatocytes. Data from this study demonstrate that diet-induced changes in the activities of promoter 1 and promoter 2 in livers of intact chicks are mimicked in chick embryo hepatocyte cultures by manipulating the concentrations of T3, glucagon and medium-chain fatty acids in the culture medium and that cis-acting sequences mediating the effects of nutrients and hormones on promoter 2 activity are located immediately upstream of the transcription start site of this promoter.
高碳水化合物喂养和三碘甲状腺原氨酸（T3）可增加禽类肝细胞中乙酰辅酶A羧化酶α（ACCα）mRNA的丰度，而饥饿、胰高血糖素和中链脂肪酸则会降低ACCα mRNA的丰度。ACCα mRNA水平的这些变化是由ACCα基因转录速率的改变介导的。在肝脏中，ACCα转录从两个启动子，即启动子1和启动子2开始，产生在其5'非翻译区具有异质性的转录本。在这里，我们通过使用核糖核酸酶保护试验测量从启动子1和启动子2表达的转录本水平，研究了启动子1和启动子2在介导营养和激素诱导的ACCα mRNA丰度变化中的作用。结果表明，启动子1和启动子2在完整雏鸡肝脏中受饥饿/再喂养调节，在鸡胚肝细胞培养物中受T3、胰高血糖素和中链脂肪酸调节，并且启动子2活性的改变在营养和激素处理引起的总ACCα mRNA丰度变化中占更大比例。在完整雏鸡肝脏和鸡胚肝细胞培养物中，相对于启动子2的转录起始位点，还在-500至+1 bp之间鉴定出五个DNase超敏位点。在瞬时转染分析中，该DNase超敏区域赋予了鸡胚肝细胞中T3、胰高血糖素和中链脂肪酸对转录的调节作用。这项研究的数据表明，通过操纵培养基中T3、胰高血糖素和中链脂肪酸的浓度，在鸡胚肝细胞培养物中模拟了完整雏鸡肝脏中饮食诱导的启动子1和启动子2活性变化，并且介导营养和激素对启动子2活性影响的顺式作用序列位于该启动子转录起始位点的紧邻上游。
