Transcription regulation of mu opioid receptor gene

• Main Authors: Po-Wei Lee, 李博偉
• Other Authors: Yu-May Lee
• Format: Others
• Language: en_US
• Published: 2003
• Online Access: http://ndltd.ncl.edu.tw/handle/70361604287143672504

碩士 === 國立臺灣大學 === 生化科學研究所 === 91 === The tolerance to opioids is determined by agonist- and cAMP-dependent regulation of opioid receptor; however, the signaling and the cis-acting elements involved are not clearly defined. In this study, we demonstrated that μ-opioid receptor (MOR) gene transcription was induced in rat pheochromocytoma (PC12) cells by the treatment with fentanyl, a morphine analogue, and that the effect of fentanyl was abolished by PKA inhibitor H89, which indicated the involvement of the cAMP-dependent pathway. In the transfection experiment, the 4.7 kb MOR promoter could be induced by forskolin, and we defined the responsive element as a CRE located at —106/-111 in 5’-UTR of the MOR gene. In the result of MOR cotransfection, we also found that fentanyl can activate 4.7kb MOR promoter activity through the responsive element. Using heterologous promoter, this element could also be induced by forskolin. In EMSA, CREB was found in the protein-DNA complexes formed on the CRE box. CREB was phosphorylated in the induction, and the phosphorylation was suppressed by H89. Direct in vivo evidence for the role of CREB in the induction of MOR gene came from the evidence that a dominant- negative mutant CREB, CREB-S133A, abolished the forskolin-mediated MOR induction. Both CREB and CBP bound to this CRE-containing region of the endogenous MOR promoter were induced by forskolin in CHIP. This CRE box is conserved in mouse, rat and human MOR genes. In conclusion, this study demonstrated that opioid-triggered MOR gene induction was mediated by the sequential activation of cAMP, PKA, CREB, and the binding of CREB and CBP to MOR promoter. These findings will be very helpful for the understanding of morphine tolerance. Furthermore, we also found that the single nucleotide difference in the MOR 5’UTR region may play a potential role in the μ-opioid receptor mRNA expression of the C57BL/6By (B6) and CXBK mice. This result suggests that the 5’UTR region of the MOR gene may affect the sensitivity to opioids between these two mouse strains by the regulation of the MOR mRNA expression.