Hayden J M, Marten N W, Burke E J, Straus D S
Biomedical Sciences Division, University of California, Riverside 92521-0121.
Endocrinology. 1994 Feb;134(2):760-8. doi: 10.1210/endo.134.2.8299571.
The abundance of insulin-like growth factor I (IGF-I) messenger RNA (mRNA) is decreased in the liver of fasting, protein-restricted, and energy-restricted rats. The extent to which this decrease in steady state mRNA abundance may be attributed to a decrease in IGF-I gene transcription remains unresolved. In the present study, we used an RNase protection assay to quantify IGF-I nuclear transcript (pre-mRNA) and mRNA abundance in whole cellular RNA isolated from liver of fasted and nonfasted male rats (4-6 weeks of age). The results of the RNase protection assay of IGF-I nuclear transcripts were strongly correlated with the results of nuclear transcription elongation (run-on) assays (r > 0.90; P < 0.001). In addition, the RNase protection assay allows for a greater capability for sensitively monitoring gene transcription in a large number of samples. In four different experiments, a consistent decrease in the quantity of IGF-I nuclear transcripts was observed in liver of animals fasted for 72 h, whereas IGF-I pre-mRNA abundance in animals fed ad libitum was highly variable (average intraassay coefficient of variation = 74% vs. 34% for nonfasted and fasted groups). When data from the four experiments were pooled, fasting reduced IGF-I pre-mRNA and mRNA levels by 78% and 70% (P < 0.001), respectively. Fasting also caused a significant decrease in mRNA and nuclear transcript abundance for another nutritionally sensitive gene, the gene encoding transthyretin (TTR). To determine whether the decrease in IGF-I and TTR nuclear transcripts was gene specific, levels of nuclear transcripts for serum albumin, H-ferritin, and ribosomal RNA were also quantified. The results indicated that serum albumin, H-ferritin, and ribosomal RNA nuclear transcripts were not decreased by fasting, demonstrating that the negative effect of fasting was specific for IGF-I and TTR. In summary, these results indicate that IGF-I and TTR nuclear transcripts are specifically decreased by fasting. The decrease in IGF-I mRNA is matched by a similar decrease in IGF-I nuclear transcripts, suggesting that fasting controls IGF-I gene expression primarily at the transcriptional level.
在禁食、蛋白质限制和能量限制的大鼠肝脏中，胰岛素样生长因子I（IGF-I）信使核糖核酸（mRNA）的丰度降低。稳态mRNA丰度的这种降低在多大程度上可归因于IGF-I基因转录的减少仍未解决。在本研究中，我们使用核糖核酸酶保护试验来定量从禁食和未禁食的雄性大鼠（4 - 6周龄）肝脏中分离的全细胞RNA中的IGF-I核转录本（前体mRNA）和mRNA丰度。IGF-I核转录本的核糖核酸酶保护试验结果与核转录延伸（连续转录）试验结果高度相关（r > 0.90；P < 0.001）。此外，核糖核酸酶保护试验在灵敏监测大量样本中的基因转录方面具有更强的能力。在四个不同的实验中，观察到禁食72小时的动物肝脏中IGF-I核转录本的数量持续减少，而随意进食动物的IGF-I前体mRNA丰度变化很大（非禁食组和禁食组的平均试验内变异系数分别为74%和34%）。当将四个实验的数据汇总时，禁食分别使IGF-I前体mRNA和mRNA水平降低了78%和70%（P < 0.001）。禁食还导致另一个营养敏感基因——编码甲状腺素转运蛋白（TTR）的基因的mRNA和核转录本丰度显著降低。为了确定IGF-I和TTR核转录本的减少是否具有基因特异性，还对血清白蛋白、H-铁蛋白和核糖体RNA的核转录本水平进行了定量。结果表明，禁食并未使血清白蛋白、H-铁蛋白和核糖体RNA的核转录本减少，这表明禁食的负面影响对IGF-I和TTR具有特异性。总之，这些结果表明禁食会使IGF-I和TTR核转录本特异性减少。IGF-I mRNA的减少与IGF-I核转录本的类似减少相匹配，这表明禁食主要在转录水平上控制IGF-I基因表达。
