Neuron-restrictive silencer factor (NRSF) functions as a repressor in neuronal cells to regulate the mu opioid receptor gene.
The mu opioid receptor (MOR) is expressed in the central nervous system and specific cell lines with varying expression levels perhaps playing important roles. One of the neuronal-specific transcription regulators, neuron-restrictive silencer factor (NRSF), has been shown to repress the expression of neuron-specific genes in non-neuronal cells. However, we showed here that the neuron-restrictive silencer element (NRSE) of MOR functions as a critical regulator to repress the MOR gene expression in specific neuronal cells depending on NRSF expression level. Using co-transfection studies, we showed that the NRSE of the MOR promoter is functional in NRSF-positive cells (NS20Y and HeLa) but not in NRSF-negative cells (PC12). NRSF binds to the NRSE of the MOR gene in a sequence-specific manner confirmed by supershift and chromatin immunoprecipitation assays, respectively. The suppression of NRSF activity with either trichostatin A or a dominant-negative NRSF induced MOR promoter activity and transcription of the MOR gene. When the NRSF was disrupted in NS20Y and HeLa cells using small interfering RNA, the transcription of the endogenous target MOR gene increased significantly. This provides direct evidence the role of NRSF in the cells and also indicates that NRSF expression is regulated by post-translational modification in neuronal NMB cells. Our data suggested that NRSF can function as a repressor of MOR transcription in specific cells, via a mechanism dependent on the MOR NRSE.
Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA. kimxx313@umn.edu
Chun Sung
CS
Kim
Cheol Kyu
CK
Hwang
Hack Sun
HS
Choi
Kyu Young
KY
Song
Ping-Yee
PY
Law
Li-Na
LN
Wei
Horace H
HH
Loh
D000818:Animals; D001483:Base Sequence; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism*; D016716:PC12 Cells; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011499:Protein Processing, Post-Translational; D012313:RNA / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017450:Receptors, Opioid, mu / Q000235:genetics*; D012097:Repressor Proteins / Q000378:metabolism / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic; D014162:Transfection; D000099229:RE1-Silencing Transcription Factor
D002843:Chromatin; D008156:Luciferases; D012313:RNA; D034741:RNA, Small Interfering; D017450:Receptors, Opioid, mu; D012097:Repressor Proteins; D014157:Transcription Factors; D000099229:RE1-Silencing Transcription Factor