Translational repression of mouse mu opioid receptor expression via leaky scanning.
Mu opioid receptor (MOR) expression is under temporal and spatial controls, but expression levels of the MOR gene are relatively low in vivo. In addition to transcriptional regulations, upstream AUGs (uAUGs) and open reading frames (uORFs) profoundly affect the translation of the primary ORF and thus the protein levels in several genes. The 5'-untranslated region (UTR) of mouse MOR mRNA contains three uORFs preceding the MOR main initiation codon. In MOR-fused EGFP or MOR promoter/luciferase reporter constructs, mutating each uAUG individually or in combinations increased MOR transient heterologous expression in neuroblastoma NMB and HEK293 cells significantly. Translation of such constructs increased up to 3-fold without altering the mRNA levels if either the third uAUG or both the second and third AUGs were mutated. Additionally, these uAUG-mediated translational inhibitions were independent of their peptide as confirmed by internal mutation analyses in each uORF. Translational studies indicated that protein syntheses were initiated at these uAUG initiation sites, with the third uAUG initiating the highest translation level. These results support the hypothesis that uORFs in mouse MOR mRNA act as negative regulators through a ribosome leaky scanning mechanism. Such leaky scanning resulted in the suppression of mouse MOR under normal conditions.
Song
Kyu Young
KY
Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA. songx047@umn.edu
Hwang
Cheol Kyu
CK
Kim
Chun Sung
CS
Choi
Hack Sun
HS
Law
Ping-Yee
PY
Wei
Li-Na
LN
Loh
Horace H
HH
D020121:5' Untranslated Regions / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D018387:Codon, Initiator; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D014176:Protein Biosynthesis; D017450:Receptors, Opioid, mu / Q000096:biosynthesis / Q000235:genetics*; D012270:Ribosomes / Q000378:metabolism
D020121:5' Untranslated Regions; D018387:Codon, Initiator; C404943:Oprm protein, mouse; D017450:Receptors, Opioid, mu