Yaffe B M, Samuels H H
J Biol Chem. 1984 May 25;259(10):6284-91.
Using cultured GH1 cells, we reported that stimulation (3- to 5-fold) of growth hormone synthesis and mRNA levels by thyroid hormone is mediated by a chromatin-associated receptor. Thyroid hormone also elicits a rapid reduction of homologous receptor in GH1 cells primarily by decreasing the synthetic rate of receptor ( Raaka , B. M., and Samuels , H. H. (1981) J. Biol. Chem. 256, 6883-6889). Without 3,5,3'-triiodo-L-thyronine (L-T3), glucocorticoid agonists induced a limited and delayed effect while L-T3 + glucocorticoid synergistically stimulated the response an additional 2- to 4-fold compared to L-T3. In this study, we utilized GC cells, a related cell line, to compare the abundance of L-T3-receptor complexes to the rate of growth hormone mRNA synthesis and gene transcription. Gene transcription was assessed by in vitro labeling of nuclei with [alpha-32P]UTP which were derived from cells incubated with hormone(s), while mRNA synthesis was determined in intact cells by [3H]uridine labeling. Labeled growth hormone mRNA and gene transcripts were quantitated by filter hybridization to plasmid containing growth hormone cDNA. L-T3 rapidly decreased receptor levels in GC cells with kinetics similar to that in GH1 cells. Both the L-T3 and the synergistic L-T3 + glucocorticoid stimulation of growth hormone mRNA synthesis changed in parallel with the level of L-T3-receptor complexes. Glucocorticoid hormones alone elicited a variable response which resulted in minimal stimulation or inhibition of growth hormone mRNA synthesis or gene transcription rates. No apparent lag was identified between the kinetics of L-T3 binding to receptor and stimulation of growth hormone gene transcription. L-T3 stimulated growth hormone gene transcription rates maximally in 1 h which then progressively decreased in parallel with L-T3-receptor levels. Using [3H]uridine pulse-chase, growth hormone mRNA was found to have a half-life of approximately 50 h in agreement with the decay curve of growth hormone production of deinduced cells. Our studies suggest that regulation of the growth hormone response is predominantly determined by positive control of growth hormone gene transcription which is proportional to the concentration of thyroid hormone-receptor complexes.
我们曾报道，利用培养的GH1细胞，甲状腺激素对生长激素合成及mRNA水平的刺激作用（3至5倍）是由一种与染色质相关的受体介导的。甲状腺激素还主要通过降低受体的合成速率，使GH1细胞中同源受体迅速减少（拉卡，B.M.，以及塞缪尔斯，H.H.（1981年）《生物化学杂志》256卷，6883 - 6889页）。在没有3,5,3'-三碘-L-甲状腺原氨酸（L-T3）的情况下，糖皮质激素激动剂产生的作用有限且延迟，而与L-T3相比，L-T3 + 糖皮质激素协同刺激反应增强了2至4倍。在本研究中，我们利用GC细胞（一种相关细胞系）来比较L-T3-受体复合物的丰度与生长激素mRNA合成及基因转录速率。通过用[α-32P]UTP对来自用激素孵育的细胞的细胞核进行体外标记来评估基因转录，而通过[3H]尿苷标记完整细胞来测定mRNA合成。通过与含有生长激素cDNA的质粒进行滤膜杂交来定量标记的生长激素mRNA和基因转录本。L-T3使GC细胞中的受体水平迅速降低，其动力学与GH1细胞中的相似。L-T3以及L-T3 + 糖皮质激素对生长激素mRNA合成的协同刺激作用均与L-T3-受体复合物的水平平行变化。单独的糖皮质激素产生的反应各不相同，导致对生长激素mRNA合成或基因转录速率的刺激作用最小或产生抑制。在L-T3与受体结合的动力学和生长激素基因转录的刺激之间未发现明显延迟。L-T3在1小时内最大程度地刺激生长激素基因转录速率，随后随着L-T3-受体水平的下降而逐渐降低。使用[3H]尿苷脉冲追踪法，发现生长激素mRNA的半衰期约为50小时，这与去诱导细胞中生长激素产生的衰减曲线一致。我们的研究表明，生长激素反应的调节主要由生长激素基因转录的正调控决定，而这与甲状腺激素-受体复合物的浓度成正比。
