|Budget Amount *help
¥3,900,000 (Direct Cost : ¥3,900,000)
Fiscal Year 1998 : ¥1,600,000 (Direct Cost : ¥1,600,000)
Fiscal Year 1997 : ¥2,300,000 (Direct Cost : ¥2,300,000)
Glucocorticoids perform these functions by binding to a cytoplasmic receptor protein glucocorticoid receptor (GR), which is a member of the nuclear receptor superfamily and acts as a ligand-inducible transcription factor. UDCA, which has widely been used as a therapeutic drug for patients with cholesterol gall stones, is currently indicated for various liver diseases including primary biliary cirrhosis, primary scierosing cholangitis, and viral hepatitis. We showed that UDCA-dependent promotion of translocation of the GR may be associated with, at least in part, its immunomodulatory action through GR-mediated gene regulation. On the other hand, nuclear import is an essential step in transcriptional regulation of gene expression by the GR.Green fluorescent protein (GFP) has been developed as a protein tag for in situ and real time visualization of desired proteins, and subcellular localization of the GR has been studied in living cells using GFP-GR chimera. We confirmed that GFP-GR dock
s in the cytoplasm in the absence of hormonal ligands and that the translocation of GFP-GR is strictly controlled in a ligand-dependent fashion. The kinetics of GFP-GR translocation appears to be similar to those reported in native GR.Since not only nuclear transport but also transactivational potential of GFP-GR is comparable to those of native GR, the translocation assay system using GFP-GR appears to be suitable for analyzing effects of various ligands and pharmacological reagents on glucocorticoid signal transduction pathway. Indeed, using this GFP-GR system, we have already shown that nuclear import of the GR is redox-sensitive. This system may also be applicable to clinical specimens to analyze tissue- or cell-type specific glucocorticoid responsiveness. Therefore, we are planning to initially screen thousands of compounds in terms of promotion of nuclear translocation of the GR using this GFP-GR system.
生細胞を用いてgreen fluorescent proteinで標識したグ受容体の核移行を検出するシステムを構築した。かかるシステムは他の転写因子の機能解析にも応用可能であることを証明した。