Identification and functional significance of polymorphisms in the mu-opioid receptor gene (Oprm) promoter of C57BL/6 and DBA/2 mice.
C57BL/6J and DBA/2J mice demonstrate differences in morphine preference when tested in a two-bottle choice paradigm. Quantitative trait loci (QTL) mapping suggested the proximal region of chromosome 10 was responsible for 41% of the observed genetic variance. The mu-opioid receptor (MOR) gene (Oprm) maps to this region and is a prime candidate for explaining the QTL. We hypothesized that variations in Oprm between these strains are responsible for differences in morphine preference. We identify five single nucleotide polymorphisms (SNPs) in the Oprm promoter; three within or near putative transcription factor binding sites. Promoter fragments were amplified from genomic DNA by polymerase chain reaction (PCR) and subcloned into luciferase reporter vectors. A significant difference in basal Oprm promoter activity was seen with C57BL/6 and DBA/2 approximately 1675 constructs in MOR-positive BE(2)-C cells, but not in MOR-negative Neuro-2a cells. In BE(2)-C cells, average DBA/2 approximately 1675 construct activity was 1.3-2.0x greater than average C57BL/6 activity suggesting that the SNPs might alter MOR expression in these two mouse strains. Significant differences in promoter activities between the two cell lines suggest that cell-type-specific transcription factors are involved. No significant differences in construct activity were found between untreated and morphine-treated BE(2)-C or Neuro-2a cells, suggesting that morphine does not regulate transcription of Oprm.
Doyle
Glenn A
GA
The Center for Neurobiology and Behavior, Department of Psychiatry, University of Pennsylvania, Philadelphia, PA 19104, United States. e_gad@verizon.net
Sheng
X Rebecca
XR
Schwebel
Candice L
CL
Ferraro
Thomas N
TN
Berrettini
Wade H
WH
Buono
Russell J
RJ
D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism / Q000503:physiopathology; D001923:Brain Chemistry / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D020022:Genetic Predisposition to Disease / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008969:Molecular Sequence Data; D009020:Morphine / Q000494:pharmacology; D009021:Morphine Dependence / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D009294:Narcotics / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D017450:Receptors, Opioid, mu / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity
D009294:Narcotics; D017450:Receptors, Opioid, mu; D009020:Morphine