The regulation of the synthesis and secretion of apolipoprotein E (apoE) is incompletely understood. This study examines the mechanisms responsible for regulating apoE gene expression in HepG, cells by thyroid hormone (3,3'-5-triiodothyronine). The secretion rate of apoE was by thyroid hormone increased (1.5 -1 %fold) in pulsekhase experiments. Thyroid hormone doubled apoE mRNA concentration as determined by Northern-blot analysis. Inhibition of protein synthesis by cycloheximide increased the thyroid-hormone-induced stimulation of apoE mRNA. This suggests that the synthesis of new protein is not required for thyroid hormone to stimulate apoE mRNA. Actinomycin D was used to inhibit new transcription; there was a more rapid degradation of mature apoE mRNA in thyroid hormone-treated HepG, cells than in control cells, suggesting that thyroid hormone acts post-transcriptionally to regulate apoE gene expression. Cycloheximide blocked the action of thyroid hormone, suggesting that thyroid hormone regulates the turnover of apoE mRNA via the synthesis of de novo protein. Nuclear run-on transcription assays demonstrated that thyroid hormone stimulated apoE gene transcription threefold in 24 h. These findings indicate that the expression of the apoE gene is controlled at both transcriptional and post-transcriptional loci by the thyroid hormone.Apolipoprotein E (apoE) is a major component of several classes of lipoproteins and plays an important role in cholesterol metabolism. Its principal function is the mediation of the cellular uptake of lipids by serving as a ligand for the apoB,E (low-density lipoprotein) receptor and for the remnant receptor (for review, see [l]). Although the liver appears to be the major site of apoE synthesis, a wide variety of peripheral tissues also produce this protein, in contrast to most apolipoproteins [2 -51.Several dietary and hormonal disturbances have been used to gain insight into the regulation of hepatic apolipoprotein synthesis, transport, lipid association and the secretion of lipoprotein particles. Hormonal stimuli affecting hepatic apolipoprotein synthesis by the rat liver in vivo include insulin 161, corticosteroid [7], sex steroids [8], and thyroid hormones [9, 101. When injected into living rats, thyroid hormones induced an increase in hepatic apoAI mRNA 19, 11, 121 and, to a lesser extent, in apoAIV mRNA [9, 111. The change in the transcription rates of apoAI and apoAIV paralleled those of their mRNA concentrations [ll]. However, Strobl et al. found that the major effect of thyroid hormones on apoAI gene expression was post-transcriptional, leading to increased stability of nuclear apoAI mRNA 1131. The synthesis of apoB48 by hyperthyroid rat liver was also greater than in hypothyroid rat liver, at the expense of apoBlOO due to an enhanced post-transcriptional introduction of a stop codon into apoB mRNA [14].The expression of the apoE gene in the liver is known to be increased in vivo by fasting [15], a sucrose diet [16], injection of glucagon or CAMP [17], but is inhibited by...

Hyperthyroidism is associated with elevated plasma levels of apolipoprotein AI (apo AI). We have examined the effects of 3,3',-5-triiodothyronine on apo AI mRNA, transcription run-on activity, apo AI mRNA half-life, and the rate of protein synthesis in Hep G2 cells, to understand the molecular mechanism by which thyroid hormone regulates apo AI gene expression. Incubation with thyroid hormone increased the apo AI and apo AII mRNA concentrations twofold. Cycloheximide alone caused a significant increase in apo AI mRNA. Nuclear run-on assays indicate that thyroid hormone did not change the rate of the apo AI gene transcription at 6, 12 or 24 h, showing that thyroid hormone did not modulate apo AI gene transcription. Kinetic studies performed in the presence of actinomycin D showed that the half-life of apo AI mRNA was increased 2-3-fold by thyroid hormone over control cells. Thyroid hormone did not change the incorporation of [35S]methionine into immunoprecipitable apo AI. Pulse-chase experiments demonstrated that there was no change in the secretion and degradation rates of labeled apo AI in response to T3. This suggests that thyroid hormone does not affect the catabolism of apo AI (degradation or/and uptake) and that translation control strongly influences the regulation of apo AI gene expression. The stabilization of apo AI mRNA by thyroid hormone and its role in translation remain to be elucidated.

No abstract

Hyperthyroidism is associated with elevated plasma levels of apolipoprotein AI (apo AI). We have examined the effects of 3,3′,‐5‐triiodothyronine on apo AI mRNA, transcription run‐on activity, apo AI mRNA half‐life, and the rate of protein synthesis in Hep G2 cells, to understand the molecular mechanism by which thyroid hormone regulates apo AI gene expression. Incubation with thyroid hormone increased the apo AI and apo AII mRNA concentrations twofold. Cycloheximide alone caused a significant increase in apo AI mRNA. Nuclear run‐on assays indicate that thyroid hormone did not change the rate of the apo AI gene transcription at 6, 12 or 24 h, showing that thyroid hormone did not modulate apo AI gene transcription. Kinetic studies performed in the presence of actinomycin D showed that the half‐life of apo AI mRNA was increased 2–3‐fold by thyroid hormone over control cells. Thyroid hormone did not change the incorporation of [35S]methionine into immunoprecipitable apo AI. Pulse‐chase experiments demonstrated that there was no change in the secretion and degradation rates of labeled apo AI in response to T3. This suggests that thyroid hormone does not affect the catabolism of apo AI (degradation or/and uptake) and that translation control strongly influences the regulation of apo AI gene expression. The stabilization of apo AI mRNA by thyroid hormone and its role in translation remain to be elucidated.

No abstract