Related Experiment Video
Updated: May 23, 2026
Reverse Yeast Two-hybrid System to Identify Mammalian Nuclear Receptor Residues that Interact with Ligands and/or Antagonists
Published on: November 15, 2013
Establishment of a cell-based drug screening model for identifying agonists of human peroxisome
Jing-Jing Ma1, Tao Zhang, Ning Fang
1Key Laboratory of Cell Engineering of Guizhou Province, the Affiliated Hospital of Zunyi Medical College, China.
Objectives:
Peroxisome proliferator-activated receptor gamma (PPARγ) plays a critical role in regulation of diverse biological processes, including lipid metabolism and adipogenesis, cell division and apoptosis, and is involved in variety of disease conditions, such as obesity, atherosclerosis, inflammation and tumour. Developing a cell-based reporter gene model targeting PPARγ would be useful to screen human PPARγ agonists that could be beneficial to patients with these diseases.
Methods:
We stably co-transfected human embryonic kidney (HEK) cell line 293T cells with phPPARγ-IRES2-EGFP vector to express human PPARγ (hPPARγ), a reporter vector pPPRE×3-TK-LUC, and control vector pRL-CMV. The efficiency of the co-transfection was evaluated with flow cytometry of hPPARγ expressing cells. Specificity of hPPARγ activity was determined by dual luciferase reporter assay of co-transfected cells exposed to PPARγ agonist rosiglitazone, PPARα agonist WY14643 and retinoic acid receptor alpha (RARα) agonist all-trans-retinoic acid (ATRA).
Key Findings:
The phPPARγ-IRES2-EGFP co-transfected HEK293T cells showed concentration- and time-dependent luciferase induction upon exposure to the rosiglitazone, while WY14643 and ATRA were unable to activate the co-transfected HEK293T cells.
Conclusions:
These data indicated that the HEK293T cells could be stably transfected with hPPARγ. This cell-based drug screening platform could be used targeting specific nuclear receptor of hPPARγ with effectiveness and specificity for hPPARγ agonists discovery.
Insights
Researchers developed a new cell model to screen for drugs targeting peroxisome proliferator-activated receptor gamma (PPARγ). This model successfully identified specific PPARγ agonists, aiding in the discovery of potential treatments for diseases like obesity and cancer.
Area of Science:
- Molecular biology
- Cell biology
- Pharmacology
Background:
- Peroxisome proliferator-activated receptor gamma (PPARγ) is crucial for regulating lipid metabolism, adipogenesis, cell division, and apoptosis.
- Dysregulation of PPARγ is implicated in diseases such as obesity, atherosclerosis, inflammation, and cancer.
Purpose of the Study:
- To develop a stable, cell-based reporter gene model for screening human PPARγ (hPPARγ) agonists.
- To create a platform for discovering novel therapeutic agents targeting hPPARγ.
Main Methods:
- Human embryonic kidney (HEK) 293T cells were stably co-transfected with vectors to express hPPARγ and a luciferase reporter.
- Reporter gene activity was assessed using a dual luciferase assay.
- Specificity was tested against agonists for PPARγ, PPARα, and RARα.
Main Results:
- The engineered HEK293T cells exhibited concentration- and time-dependent luciferase induction in response to rosiglitazone, a known PPARγ agonist.
- The reporter system did not respond to PPARα or RARα agonists, confirming specificity.
Conclusions:
- Stable transfection of HEK293T cells with hPPARγ and a reporter system is feasible.
- This cell-based platform offers an effective and specific method for discovering hPPARγ agonists.