The mechanism of glucocorticoid receptor dependent regulation of NRF2 activity
- 주제(키워드) NRF2 , glucocorticoid
- 주제(DDC) 615.1
- 발행기관 아주대학교
- 지도교수 전상민
- 발행년도 2022
- 학위수여년월 2022. 2
- 학위명 석사
- 학과 및 전공 일반대학원 약학과
- 실제URI http://www.dcollection.net/handler/ajou/000000031751
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Nuclear factor E2-related factor 2 (NRF2) plays a key role in redox metabolism and antioxidant defense. Recent studies showed that abnormal activation of NRF2 is a common event in many types of cancer suggesting that NRF2 could be a promosing target for cancer therapy. Previously, we identified clobetasol propionate (CP), a glucocorticoid analogue as the most potent NRF2 inhibitor from a clinical compound library screening. In addition, we also reported that CP inhibits NRF2 through glucocorticoid receptor (GCR) and β-TrCP -dependent proteasomal degradation of NRF2. In the present study, we further investigated the mechanisms of CP on NRF2 inhibition and the crosstalk between GR and NRF2. Either inhibition of protein synthesis by cycloheximide (CHX) or CP rapidly induced degradation of NRF2 protein in 1hr or 2hr, respectively. In addition, NRF2 degradation was further accelerated in the presence of both CHX and CP, supporting the finding that CP promotes NRF2 protein degradation. Using the DNA non-binding mutants of GCR, we also confirmed that the effect of CP on NRF2 inhibition is independent on the transcriptional activity, rather dependent on non-genomic activity of GCR. Interestingly, the non-genomic mechanism is also supported by the finding that XA compound, which we identified from the screening, suppressed CP-induced GCR transcriptional activity without compromising CP-induced NRF2 inhibition. Indeed, we confirmed that GCR co-immunoprecipitaed with NRF2. Finally, we also found regulatory crosstalk between GCR and NRF2, which GCR can promote the expression and activity of NRF2 while NRF2 can suppress GCR expression and activity. Taken together, our data demonstrate the novel mechanisms of GCR-dependent NRF2 inhibition and their cross-regulatory network.
more목차
Ⅰ. INTRODUCTION 1
Ⅱ. MATERIALS AND METHODS 4
A. Cell culture and Reagents 4
B. Luciferase assay 4
C. Western blot 5
D. Immunostaining 5
E. Fractionation 5
F. Co-IP 6
Ⅲ. PRELIMINARY RESEARCH 7
Ⅳ. RESULTS 9
A. Effect of glucocorticoid on the stability of NRF2 9
B. The NRF2 inhibitory effect of CP does not require DNA binding 11
C. Confirmation of binding of GCR and NRF2 during CP treatment 16
D. Effect of GCR on the stability of NRF2 17
E. Effect of GCR on the expression of NRF2 21
Ⅴ. DISCUSSION AND CONCLUSION 24
Ⅵ. REFERENCE 25
국문요약 28
