Evidence of endogenous mu opioid receptor regulation by epigenetic control of the promoters.
The pharmacological effect of morphine as a painkiller is mediated mainly via the mu opioid receptor (MOR) and is dependent on the number of MORs in the cell surface membrane. While several studies have reported that the MOR gene is regulated by various cis- and trans-acting factors, many questions remain unanswered regarding in vivo regulation. The present study shows that epigenetic silencing and activation of the MOR gene are achieved through coordinated regulation at both the histone and DNA levels. In P19 mouse embryonal carcinoma cells, expression of the MOR was greatly increased after neuronal differentiation. MOR expression could also be induced by a demethylating agent (5'-aza-2'-deoxycytidine) or histone deacetylase inhibitors in the P19 cells, suggesting involvement of DNA methylation and histone deacetylation for MOR gene silencing. Analysis of CpG DNA methylation revealed that the proximal promoter region was unmethylated in differentiated cells compared to its hypermethylation in undifferentiated cells. In contrast, the methylation of other regions was not changed in either cell type. Similar methylation patterns were observed in the mouse brain. In vitro methylation of the MOR promoters suppressed promoter activity in the reporter assay. Upon differentiation, the in vivo interaction of MeCP2 was reduced in the MOR promoter region, coincident with histone modifications that are relevant to active transcription. When MeCP2 was disrupted using MeCP2 small interfering RNA, the endogenous MOR gene was increased. These data suggest that DNA methylation is closely linked to the MeCP2-mediated chromatin structure of the MOR gene. Here, we propose that an epigenetic mechanism consisting of DNA methylation and chromatin modification underlies the cell stage-specific mechanism of MOR gene expression.
Hwang
Cheol Kyu
CK
Department of Pharmacology, University of Minnesota, 6-120 Jackson Hall, 321 Church St. S.E., Minneapolis, MN 55455, USA. hwang025@umn.edu
Song
Kyu Young
KY
Kim
Chun Sung
CS
Choi
Hack Sun
HS
Guo
Xiao-Hong
XH
Law
Ping-Yee
PY
Wei
Li-Na
LN
Loh
Horace H
HH
D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D044127:Epigenesis, Genetic / Q000187:drug effects; D006657:Histones / Q000378:metabolism; D006877:Hydroxamic Acids / Q000494:pharmacology; D051783:Methyl-CpG-Binding Protein 2 / Q000378:metabolism; D008745:Methylation / Q000187:drug effects; D051379:Mice; D008954:Models, Biological; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D034741:RNA, Small Interfering / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D006657:Histones; D006877:Hydroxamic Acids; D051783:Methyl-CpG-Binding Protein 2; D034741:RNA, Small Interfering; D017450:Receptors, Opioid, mu; C012589:trichostatin A; D001374:Azacitidine