Development and optimization of a cell screening system for farnesoid X receptor agonist.
- Author:
Zhimin ZHENG
1
;
Xiaoxia HUANG
1
;
Biying PANG
1
;
Nana HUANG
1
;
Bo KONG
1
;
Xin LI
1
;
Wenting XIONG
1
Author Information
- Publication Type:Journal Article
- Keywords: FXR response element; dual-luciferase report system; high throughput screening; nuclear receptor
- MeSH: Humans; Mice; Animals; Transcription Factors/genetics*; DNA-Binding Proteins/genetics*; Receptors, Cytoplasmic and Nuclear/genetics*; Genes, Reporter; Luciferases/genetics*
- From: Chinese Journal of Biotechnology 2023;39(1):359-371
- CountryChina
- Language:Chinese
- Abstract: This study aims to develop an improved cell screening system for farnesoid X receptor (FXR) agonists based on a dual luciferase reporter gene system. FXR response element (FXRE) fragments from FXR target genes were cloned and inserted into upstream of firefly luciferase (Luc) gene in the plasmid pGL4-luc2P-Hygro. In combination with the internal reference plasmid containing renilla luciferase, a dual luciferase reporter gene system was developed and used for high throughput screening of FXR agonists. After studying the effects of over-expression of RXR, mouse or human FXR, various FXRE fragments, and different ratio of FXR plasmid amount to reporter gene plasmid, induction efficiency of the screening system was optimized by the known FXR agonist GW4064, and Z factor for the system reached 0.83 under optimized conditions. In summary, an improved cell screening system based on double luciferase reporter gene detection system was developed to facilitate the discovery of FXR agonists, where a new enhanced FXRE element was formed by a superposition of multiple FXRE fragments from FXR target genes, instead of a superposition of traditional IR-1 (inverted repeats-1) fragments.
