Identification of Mel1a melatonin receptors in the human embryonic kidney cell line HEK293: evidence of G protein-coupled melatonin receptors which do not mediate the inhibition of stimulated cyclic AMP levels.
Binding assays using 2-[125I]iodomelatonin revealed high-affinity, guanosine 5'-O-(3-thiotriphosphate) sensitive, melatonin binding sites (B(max) 1.1 fmol/mg protein) in the human embryonic kidney cell line HEK293. Competition studies using the selective melatonin receptor antagonist luzindole and RT-PCR techniques identified these sites as human Mel1a melatonin receptors. Challenge of HEK293 cells with 1 microM melatonin had no effect on forskolin stimulated cyclic AMP levels, whereas in HEK293 cells engineered to stably over-express the human Mel1a melatonin receptor (B(max) > 400 fmol/mg protein) melatonin dose-dependently inhibited stimulated cyclic AMP levels (IC50 7.7 pM). These data may indicate that certain tissues, expressing low levels of G protein-coupled melatonin receptors, do not display melatonin mediated inhibition of cAMP.
Conway
S
S
Molecular Neuroendocrinology Group, Rowett Research Institute, Bucksburn, Aberdeen, UK. sco@rri.sari.ac.uk
Drew
J E
JE
Canning
S J
SJ
Barrett
P
P
Jockers
R
R
Strosberg
A D
AD
Guardiola-Lemaitre
B
B
Delagrange
P
P
Morgan
P J
PJ
D001667:Binding, Competitive; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D015698:Genomic Library; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D008550:Melatonin / Q000031:analogs & derivatives / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D044094:Receptors, Melatonin; D017422:Sequence Analysis, DNA; D012701:Serotonin / Q000031:analogs & derivatives / Q000378:metabolism; D015398:Signal Transduction; D014363:Tryptamines / Q000378:metabolism
D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D044094:Receptors, Melatonin; D014363:Tryptamines; C057154:luzindole; D005576:Colforsin; D012701:Serotonin; C044556:2-iodomelatonin; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D008550:Melatonin; C006389:N-acetylserotonin; C012365:6-hydroxymelatonin