Protein interactome mining defines melatonin MT1 receptors as integral component of presynaptic protein complexes of neurons

In mammals, the hormone melatonin is mainly produced by the pineal gland with nocturnal peak levels. Its peripheral and central actions rely either on its intrinsic antioxidant properties or on binding to melatonin MT 1 and MT 2 receptors, belonging to the G protein-coupled receptor (GPCR) super-family. Melatonin has been reported to be involved in many functions of the central nervous system such as circadian rhythm regulation, neurotransmission, synaptic plasticity, memory, sleep, and also in Alzheimer's disease and depression. However, little is known about the subcellular localization of melatonin receptors and the molecular aspects involved in neuronal functions of melatonin. Identification of protein complexes associated with GPCRs has been shown to be a valid approach to improve our understanding of their function. By combining proteomic and genomic approaches we built an interactome of MT 1 and MT 2 receptors, which comprises 378 individual proteins. Among the proteins interacting with MT 1 , but not with MT 2 , we identified several presynaptic proteins, suggesting a potential role of MT 1 in neurotransmission. Presynaptic localization of MT 1 receptors in the hypothalamus, striatum, and cortex was confirmed by subcellular fractionation experiments and immunofluorescence microscopy. MT 1 physically interacts with the voltage-gated calcium channel Ca v 2.2 and inhibits Ca v 2.2-promoted Ca 2+ entry in an agonist-independent manner. In conclusion, we show that MT 1 is part of the presynaptic protein network and negatively regulates Ca v 2.2 activity, providing a first hint for potential synaptic functions of MT 1 .