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Colorful Disks

Nuclear Receptor  Reporter Stable Cell Lines


Nuclear Hormone Receptors (NHRs) represent a fascinating class of ligand-binding transcription factors (TFs) crucial in regulating gene expression. With over 350 NHRs cataloged in the Protein Data Bank, their intricate role in cellular processes is underscored by the modulation of activities through specific ligand interactions. While conventional DNA regulatory element-luciferase reporter cell lines have been instrumental in studying these receptors, their limitations arise from a lack of specificity due to shared binding sequences among family members. Recognizing this challenge, Signosis has innovatively devised ligand-binding domain (LBD)-driven GAL4 reporter stable cell lines, a method poised to offer heightened sensitivity, specificity, and reduced toxicity. This transformative approach opens avenues for studying NHRs in a more nuanced and precise manner, particularly in drug compound screening for agonist or antagonist properties.

Principle

Nuclear hormone receptors (NHRs) constitute a group of ligand-binding transcription factors (TFs). More than 350 NHRs are available in the Protein Data Bank. Like other TFs, they can regulate gene expression by binding to specific DNA regulatory elements, but their activities are modulated by the corresponding ligands.

Although DNA regulatory element-luciferase reporter cell lines can be used to study endogenous or exogenous receptors and analyze the receptor signaling pathway in a native biological context, the NHR DNA element-luciferase reporter cell lines lack specificity and uniqueness due to the similarity in binding sequences among their entire family members.

 

Signosis has developed ligand-binding domain (LBD)-driven GAL4 reporter stable cell lines by delivering two vectors, one expressing 8 copies of GAL4 upstream activator sequences (UAS) in front of the luciferase reporter gene, and another expressing Gal4 DNA Binding Domain (DBD) bound to a ligand binding domain (LBD) of your choice. Upon addition of a corresponding ligand, the DBD-LBD fusion protein is activated, binding to the GAL promoter binding site that will drive forward the expression of luciferase. With this method, our GAL4-based cell line assays will have minimal cross-reactivity with other nuclear receptors, leading to high sensitivity and specificity, while also having low toxicity of the chimeric receptors to the cells when overexpressed. These cell lines can be used to screen drug compounds for agonist or antagonist.

Stable clones were selected with both hygromycin and G418, and a functional assay was subsequently conducted, with clones exhibiting the highest induction fold chosen.

Benefits

Highly Specific

The responsiveness of this stable cell line is driven by the specific ligand-binding domain with low/no cross-reactivity with other members of the nuclear receptor.

Routine Mycoplasma Testing

All cell lines tested negative for mycoplasma.

Less Toxic