The end of a myth: cloning and characterization of the ovine melatonin MT2 receptor
The end of a myth: cloning and characterization of the ovine melatonin MT2 receptor
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DOI：
10.1111/j.1476-5381.2009.00453.x
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发表时间：
2009-11-01
影响因子：
7.3
通讯作者：
Boutin, J. A.
中科院分区：
文献类型：
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作者：
Coge, F.;Guenin, S. P.;Boutin, J. A.
背景和目的：多年来，人们怀疑绵羊只表达一种褪黑激素受体（与其他哺乳动物的MT 1非常相似）。实验方法：利用与牛MT 2 mRNA序列同源的耐热逆转录酶和聚合酶链反应引物，从绵羊视网膜中克隆了MT 2受体，并对其进行了鉴定，结果：绵羊MT 2受体与牛、人和大鼠的同源性分别为96%、72%和67%。这种在CHO-K1细胞中稳定表达的MT 2受体显示出高亲和力的2[125 I]-碘褪黑激素结合（K-D = 0.04 nM）。褪黑素、4-苯基-2-丙酰胺四氢萘和luzindole抑制2[125 I]-碘褪黑素结合的顺序与其他物种的MT 2受体所表现出的顺序相似（褪黑素> 4-苯基-2-丙酰胺四氢萘> luzindole）。然而，其药理学特征更接近大鼠而不是人MT 2受体。在功能上，绵羊MT 2受体与G（i）蛋白偶联，导致腺苷酸环化酶的抑制，如其他褪黑激素受体一样。在绵羊脑中，MT 2 mRNA在结节部、脉络丛和视网膜中表达，在乳头体中有中等程度的表达。实时荧光定量聚合酶链反应显示，在绵羊结节部，前乳头体下丘脑和乳头体，MT 1和MT 2 mRNA的表达的时间模式是不平行的三个tissues.Conclusion和影响：MT 1和MT 2受体在所有分析的羊脑组织中的共表达表明，MT 2受体可能参与褪黑素调节季节性无排卵活动的母羊通过调制MT 1受体的行动。
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[125I]-iodomelatonin binding (K-D = 0.04 nM). The rank order of inhibition of 2[125I]-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 G(i) 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.