Establishment of reporter cells that respond to glucocorticoids by a transposon-mediated promoter-trapping system

Kosuke Ishikawa, Sakura Tamamura, Kentaro Semba, Shinya Watanabe

Research output: Contribution to journalArticlepeer-review

Abstract

Previously, we had established a highly sensitive trap vector system for the efficient isolation of reporter cells for a certain condition of interest. In this study, we used this system to screen reporter cells that express the luciferase and enhanced green fluorescent protein genes in response to dexamethasone, a glucocorticoid receptor agonist to facilitate glucocorticoid signaling research. In total, 10 clones were isolated. The insertion sites of the trap vector were analyzed using 5′ rapid amplification of cDNA ends (5′ RACE), whereupon LPIN1, PKP2, and FKBP5 were identified as genes that were upregulated by the dexamethasone treatment. Specifically, PKP2 has not previously been focused as a gene that responds to glucocorticoids. The PKP2 mRNA was analyzed and induction of the endogenous gene was confirmed by real-time polymerase chain reaction. Given that PKP2 does not appear to have a consensus glucocorticoid response element (GRE) sequence, this reporter clone could supplement the current GRE-based reporter systems that are prevalently used. Because different clones showed different responses to glucocorticoids, these clones should provide more information than analysis with a single reporter clone. This paper demonstrates that the previously developed trap vector technology can contribute to the rapid construction of drug evaluation systems.

Original languageEnglish
Article number105819
JournalEuropean Journal of Pharmaceutical Sciences
Volume162
DOIs
Publication statusPublished - 2021 Jul 1

Keywords

  • Gene reporter assay
  • Glucocorticoid receptor
  • Glucocorticoids
  • Transposon
  • Trap vector

ASJC Scopus subject areas

  • Pharmaceutical Science

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