The end of a myth: cloning and characterization of the ovine melatonin MT₂ receptor

Abstract

Background and purpose: For many years, it was suspected that sheep expressed only one melatonin receptor (closely resembling MT1 from other mammal species). Here we report the cloning of another melatonin receptor, MT2, from sheep. Experimental approach: Using a thermo-resistant reverse transcriptase and polymerase chain reaction primer set homologous to the bovine MT2 mRNA sequence, we have cloned and characterized MT2 receptors from sheep retina. Key results: The ovine MT2 receptor presents 96%, 72% and 67% identity with cattle, human and rat respectively. This MT2 receptor stably expressed in CHO-K1 cells showed high-affinity 2[ 125 I]-iodomelatonin binding (KD = 0.04 nM). The rank order of inhibition of 2[ 125 I]-iodomelatonin binding by melatonin, 4-phenyl-2-propionamidotetralin and luzindole was similar to that exhibited by MT2 receptors of other species (melatonin > 4-phenyl-2-propionamidotetralin > luzindole). However, its pharmacological profile was closer to that of rat, rather than human MT2 receptors. Functionally, the ovine MT2 receptors were coupled to Gi proteins leading to inhibition of adenylyl cyclase, as the other melatonin receptors. In sheep brain, MT2 mRNA was expressed in pars tuberalis, choroid plexus and retina, and moderately in mammillary bodies. Real-time polymerase chain reaction showed that in sheep pars tuberalis, premammillary hypothalamus and mammillary bodies, the temporal pattern of expression of MT1 and MT2 mRNA was not parallel in the three tissues. Conclusion and implications: Co-expression of MT1 and MT2 receptors in all analysed sheep brain tissues suggests that MT2 receptors may participate in melatonin regulation of seasonal anovulatory activity in ewes by modulating MT1 receptor action.

Photoperiod-dependent effect of inflammation on nocturnal gene expression of proinflammatory cytokines and their receptors in <i>pars tuberalis</i> of ewe

“…We also found that the basal gene expression of proinflammatory cytokines receptors in the PT was similar during LN and SN. The fact that LPS promotes both TNF receptors gene expression and IL6ST gene expression in LN but not in SN period may result from the different level of melatonin during these two photoperiods, because the PT is exclusive pituitary structure expressing melatonin binding sites (Cogé et al, 2009).…”

International Union of Basic and Clinical Pharmacology. LXXV. Nomenclature, Classification, and Pharmacology of G Protein-Coupled Melatonin Receptors

“…We expect the classification of melatonin receptors will continue to evolve as operational and structural data for existing receptors are further defined and possible variants and/or new receptors are characterized. This is the case for the recent cloning of the ovine MT 2 receptor, where for many years it was suspected that sheep possessed only one melatonin receptor (Cogé et al, 2009).…”

Encoding and Decoding Photoperiod in the Mammalian Pars Tuberalis

“…Although it is not functional in some seasonal species, MT2 has recently been characterised in the sheep PT, with expression levels lower than those of MT1 [118]. With its ability to form heterodimers with MT1 [117], the involvement of MT2 in melatonin signalling in the PT remains to be investigated.…”