Endocrinol Metab.  2015 Dec;30(4):584-592. 10.3803/EnM.2015.30.4.584.

Thyroid Hormone Regulates the mRNA Expression of Small Heterodimer Partner through Liver Receptor Homolog-1

• Ahn HY
• Kim HH
• Kim YA
• Kim M
• Ohn JH
• Chung SS
• Lee YK
• Park DJ
• Park KS
• Moore DD
• Park YJ

• KMID: 2169669
• DOI: http://doi.org/10.3803/EnM.2015.30.4.584

Abstract

BACKGROUND
Expression of hepatic cholesterol 7alpha-hydroxylase (CYP7A1) is negatively regulated by orphan nuclear receptor small heterodimer partner (SHP). In this study, we aimed to find whether thyroid hormone regulates SHP expression by modulating the transcriptional activities of liver receptor homolog-1 (LRH-1).
METHODS
We injected thyroid hormone (triiodothyronine, T3) to C57BL/6J wild type. RNA was isolated from mouse liver and used for microarray analysis and quantitative real-time polymerase chain reaction (PCR). Human hepatoma cell and primary hepatocytes from mouse liver were used to confirm the effect of T3 in vitro. Promoter assay and electrophoretic mobility-shift assay (EMSA) were also performed using human hepatoma cell line
RESULTS
Initial microarray results indicated that SHP expression is markedly decreased in livers of T3 treated mice. We confirmed that T3 repressed SHP expression in the liver of mice as well as in mouse primary hepatocytes and human hepatoma cells by real-time PCR analysis. LRH-1 increased the promoter activity of SHP; however, this increased activity was markedly decreased after thyroid hormone receptor beta/retinoid X receptor alpha/T3 administration. EMSA revealed that T3 inhibits specific LRH-1 DNA binding.
CONCLUSION
We found that thyroid hormone regulates the expression of SHP mRNA through interference with the transcription factor, LRH-1.

Keyword

Small heterodimer partner; Thyroid hormones; Cholesterol; Liver receptor homolog-1; Bile acids and salts

MeSH Terms

Animals
Bile Acids and Salts
Carcinoma, Hepatocellular
Cell Line
Child
Child, Orphaned
Cholesterol
Cholesterol 7-alpha-Hydroxylase
DNA
Hepatocytes
Humans
Liver*
Mice
Microarray Analysis
Real-Time Polymerase Chain Reaction
Receptors, Thyroid Hormone
RNA
RNA, Messenger*
Thyroid Gland*
Thyroid Hormones
Transcription Factors
Bile Acids and Salts
Cholesterol
Cholesterol 7-alpha-Hydroxylase
DNA
RNA
RNA, Messenger
Receptors, Thyroid Hormone
Thyroid Hormones
Transcription Factors

Figure

• 

  Fig. 1 Effect of thyroid hormone (T3) treatment on small heterodimer partner (SHP) expression. Comparison of SHP expression in mouse liver (A, n=4 or n=5 in each group) after 6 hours or 5 days of thyroid hormone treatment (1 mg/g). Comparison of SHP expression in mouse primary hepatocytes (B) and human hepatoma cells (C) after 2 and 6 hours of T3 treatment (100 nmol/L). aSignificant differences compared to the control are P<0.05.

• 

  Fig. 2 Effect of thyroid hormone (T3) on small heterodimer partner (SHP) promoter activity with or without thyroid hormone receptor (TR)/retinoid X receptor (RXR), and liver receptor homolog-1 (LRH-1). In each HepG2 cell sample, 300 ng of mouse SHP (A) or human SHP (B, C) promoter DNA was co-transfected with or without 75 ng of TRβ and 75 ng of RXRα or 75 ng of LRH-1. Vehicle or T3 (100 nmol/L) was administered for 24 hours to determine the effect of thyroid hormone on SHP promoter expression. Luciferase activity was measured and normalized to β-galactosidase activity. Significant differences compared to the control are aP<0.05 and bP<0.01.

• 

  Fig. 3 Repression of expression from the 5' deletion small heterodimer partner (SHP) promoter by the thyroid hormone. The 200 ng of 5' human SHP promoter (full, -1490 and -175) was co-transfected with or without 100 ng of thyroid hormone receptor β (TRβ) and 30 ng of retinoid X receptor α (RXRα) or 100 ng of liver receptor homolog-1 (LRH-1) in 293 cells. Thyroid hormone (T3; 100 nmol/L) was added. Luciferase activity was measured and normalized against renilla activity. NS, not significant. Significant differences compared to the control aP<0.01.

• 

  Fig. 4 Interaction between thyroid hormone (T3) and liver receptor homolog-1 (LRH-1). Electrophoretic mobility-shift assay was performed using an oligomer containing the LRH-1 responsive element (LRE) in the human SHP promoter (-518) as a probe and nuclear extracts of human hepatoma cells treated with 100 nmol/L of T3 for increasing times as indicated. For competition assays, unlabeled oligomers containing the hLRH-1(-518) LRE (self; lane 8), an LRE mutant (lane 9), or a consensus LRE (lane 10) were used in a 50-fold molar excess. MT, mutant.

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