The Modulation of Apolipoprotein E Gene Expression by 3,3′‐5‐triiodothyronine in HepG₂ Cells Occurs at Transcriptional and Post‐transcriptional Levels

Abstract

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...

Regulated expression of apolipoprotein E by human retinal pigment epithelial cells

“…LXR-RXR heterodimers, stimulated by their appropriate ligands, upregulate apoE expression (32,51,52). T 3 has also been demonstrated to increase the expression of apoE in HepG2 cells (33). Cultured human RPE cells are regulated in a similar manner.…”

“…Liver X receptor (LXR) and retinoid X receptor (RXR) ligands increase the levels of these transporters and increase reverse cholesterol transport in macrophages (30)(31)(32). Thyroid hormone (T 3 ) has also been demonstrated to increase the expression of apoE in HepG2 cells (33).…”

Apolipoprotein E is a genetic risk factor for fetal iodine deficiency disorder in China

“…However since T3 directly influences transcription of APOE, it seems equally likely that the reduced ability of ⑀4 compared to ⑀3 carriers to catalyse neuronal growth and branching of dendrites is further compromised by a change in transcription driven by reduced T3 levels. 16 Whatever the explanation a threshold leading to intellectual impairment is breached more easily with profound implications for neuronal development during key periods of fetal life.…”

“…3,15 In addition 3,5,3Ј-Triiodothyronine (T3) upregulates transcription of APOE and also acts post-transcriptionally. 16 In the light of this circumstantial evidence we decided to evaluate APOE as a candidate susceptibility locus for FIDD.…”

In vivo stimulation of the beta(2)-adrenergic pathway increases expression of the Gi proteins and the alpha(2)A-adrenergic receptor genes in the pregnant rat myometrium

“…The isolated nuclei were resuspended in 50 mM Tris-HCl (pH 8·3), 40% glycerol, 5 mM MgCl 2 and 0·1 mM EDTA and stored at 80 C. Nuclei (2 10 7 ) were incubated for 30 min at 30 C with 250 µCi [ -32 P]UTP (400 Ci/mmol) in a 200 µl mixture prepared as described by Antras et al (1991). The mixture was then treated with DNase I and proteinase K (10 mg/ ml) as previously described by Vandenbrouck et al (1994); nascent RNAs were extracted with 8 M guanidine-HCl (pH 5·0) containing 20 mM Na(CH 3 CO 2 ), 1 mM dithiothreitol and 0·5% (w/v) lauryl sarcosine. The radiolabeled RNAs (50 10 6 c.p.m.)…”