Molecular and Cellular PharmacologyDifferential signaling of human Mel1a and Mel1b melatonin receptors through the cyclic guanosine 3′-5′-monophosphate pathway
Section snippets
Cloning and expression of human Mel1a and Mel1b receptors
The human Mel1a receptor cDNA was cloned by RT-PCR from human embryonic RNA and by PCR from human genomic DNA. Human embryonic brain RNA was reverse transcripted as described [5]. Mel1a-specific primers were selected from the published sequence [8]. DNAs were amplified with high-fidelity Pfu or Pwo DNA polymerase as described previously [5]. The N-terminal part corresponding to the first exon was amplified using PCR primers 23S(5′-AACAAAGCTTATGCAGGGC-3′) and 203AS (5′-GATGTTGTTCCTGAGC)
Cloning, expression, and pharmacological characterization of human Mel1a and Mel1b receptors
The coding region and the 3′ untranslated region of the Mel1a receptor were cloned by RT-PCR from human embryonic brain RNA. Mel1a-specific PCR primers were selected from the recently published sequence [8]. The human Mel1b receptor gene was isolated from human genomic DNA by PCR using Mel1b-specific primers selected from the recently published sequence [4]. Both receptor DNAs were cloned into the expression vector pcDNA3/RSV and their identity was confirmed by sequencing. No amino acid
Discussion
Cyclic GMP is an important second messenger in the central nervous system [22], in blood vessels, and in the retina, locations where melatonin receptors are expressed 4, 6, 23, 24. Melatonin is secreted from the pineal gland in a circadian manner with maximal levels during the night. Cyclic cGMP levels have also been shown to vary in a circadian manner in the cerebral cortex of the chick, where they are correlated to blood rhythms of melatonin [25]. Interestingly, the cycle of cGMP and
Acknowledgements
This work was supported by grants from CNRS, the Association pour la Recherche sur le Cancer (N°9890), the University of Paris, and the Intitut de Recherches Internationales Servier (IRIS). We thank Drs. S. Marullo, P. J. Morgan, and P. Barret for their expert advice and help in the initial phase of receptor cloning.
References (32)
- et al.
Cloning and functional analysis of a polymorphic variant of the ovine Mel1a melatonin receptor
Biochim Biophys Acta
(1997) - et al.
Cloning and characterization of a mammalian melatonin receptor that mediates reproductive and circadian responses
Neuron
(1994) - et al.
Differential regulation of melatonin receptors in sheep, chicken and lizard brains by cholera and pertussis toxins and guanine nucleotides
Neurochem Int
(1996) - et al.
The melatonin receptor in the human brainCloning experiments and distribution studies
Brain Res Mol Brain Res
(1996) - et al.
Melatonin receptors are for the birdsMolecular analysis of two receptor subtypes differentially expressed in chick brain
Neuron
(1995) - et al.
Molecular dissection of two distinct actions of melatonin on the suprachiasmatic circadian clock
Neuron
(1997) - et al.
Melatonin receptors mediate potentiation of contractile responses to adrenergic nerve stimulation in rat caudal artery
Eur J Pharmacol
(1995) - et al.
Melatonin inhibits cyclic AMP and cyclic GMP accumulation in the rat pituitary
Brain Res
(1989) - et al.
Transduction mechanisms of vertebrate and invertebrate photoreceptors
J Biol Chem
(1994) Melatonin and 5-hydroxytryptamine phase-advance the rat circadian clock by activation of nitric oxide synthesis
Neurosci Lett
(1996)