Back D W, Wilson S B, Morris S M, Goodridge A G
J Biol Chem. 1986 Sep 25;261(27):12555-61.
Mechanisms involved in stimulation of the synthesis of malic enzyme by insulin and triiodothyronine and in inhibition of synthesis by glucagon have been investigated by assessing levels and rates of synthesis of malic enzyme mRNA in chick embryo hepatocytes in culture. Insulin alone had no effect on the level of malic enzyme mRNA, whereas triiodothyronine by itself caused a 7-fold increase. Insulin plus triiodothyronine caused an 11-fold increase. Glucagon caused a 93% decrease in the accumulation of malic enzyme mRNA caused by insulin plus triiodothyronine. Although the relative changes in mRNA level are smaller in magnitude, they are qualitatively similar to the effects of these hormones on synthesis of malic enzyme, suggesting that control is exerted primarily at pretranslational steps. After addition of triiodothyronine, malic enzyme mRNA accumulated with sigmoidal kinetics, approaching a new steady state at 36-48 h after adding hormone. Puromycin, an inhibitor of protein synthesis, blocked the effect of triiodothyronine if added 30 min prior to the hormone and inhibited further accumulation of malic enzyme mRNA if added 24 h after triiodothyronine. However, puromycin had no effect on the level of beta-tubulin mRNA (t1/2 = 3-5 h), suggesting that the effect of triiodothyronine on malic enzyme mRNA required synthesis of a peptide. Triiodothyronine increased transcription of the malic enzyme gene by 2-fold and level of its mRNA by 11-14-fold, indicating regulation is primarily at a post-transcriptional step. Glucagon caused malic enzyme mRNA to decay with a half-life of 1.5 h, whereas alpha-amanitin or actinomycin D, inhibitors of transcription, caused the mRNA to decay with a half-life of 8-11 h. The effect of glucagon was entirely post-transcriptional because the hormone had no effect on transcription. Taken together, these results suggest a model in which triiodothyronine regulates production of a peptide that stabilizes malic enzyme transcripts in the cytoplasm and/or nucleus. Glucagon may inhibit activity of the peptide induced by triiodothyronine.
通过评估培养的鸡胚肝细胞中苹果酸酶mRNA的水平和合成速率，对胰岛素和三碘甲状腺原氨酸刺激苹果酸酶合成以及胰高血糖素抑制其合成所涉及的机制进行了研究。单独使用胰岛素对苹果酸酶mRNA的水平没有影响，而三碘甲状腺原氨酸自身可使其增加7倍。胰岛素加三碘甲状腺原氨酸可使其增加11倍。胰高血糖素使胰岛素加三碘甲状腺原氨酸所引起的苹果酸酶mRNA积累减少93%。虽然mRNA水平的相对变化幅度较小，但在质量上与这些激素对苹果酸酶合成的影响相似，这表明调控主要在翻译前步骤发挥作用。加入三碘甲状腺原氨酸后，苹果酸酶mRNA呈S形动力学积累，在添加激素后36 - 48小时接近新的稳态。嘌呤霉素是一种蛋白质合成抑制剂，如果在激素添加前30分钟加入，可阻断三碘甲状腺原氨酸的作用，如果在三碘甲状腺原氨酸加入后24小时加入，则抑制苹果酸酶mRNA的进一步积累。然而，嘌呤霉素对β-微管蛋白mRNA的水平没有影响（半衰期 = 3 - 5小时），这表明三碘甲状腺原氨酸对苹果酸酶mRNA的作用需要合成一种肽。三碘甲状腺原氨酸使苹果酸酶基因的转录增加2倍，其mRNA水平增加11 - 14倍，表明调控主要在转录后步骤。胰高血糖素使苹果酸酶mRNA以1.5小时的半衰期衰减，而转录抑制剂α-鹅膏蕈碱或放线菌素D使mRNA以8 - 11小时的半衰期衰减。胰高血糖素的作用完全是转录后作用，因为该激素对转录没有影响。综上所述，这些结果提示了一个模型，其中三碘甲状腺原氨酸调节一种肽的产生，该肽可稳定细胞质和/或细胞核中的苹果酸酶转录本。胰高血糖素可能抑制三碘甲状腺原氨酸诱导的肽的活性。
