Molecular determinants for the differential coupling of Gα₁₆ to the melatonin MT1, MT2 and Xenopus Mel1c receptors

Abstract

The pineal neurohormone melatonin modulates a variety of physiological processes through different receptors. It has recently been reported that the cloned melatonin receptors (MT1, MT2 and Mel1c) exhibit differential abilities to stimulate phospholipase C (PLC) via G 16 . Here we examined the molecular basis of such differences in melatonin receptor signaling. Coexpression of MT1 or MT2 with the a subunit of G 16 (Ga 16 ) allowed COS-7 cells to accumulate inositol phosphates in response to 2-iodomelatonin. In contrast, Mel1c did not activate Ga 16 even though its expression was demonstrated by radioligand binding and agonist-induced inhibition of adenylyl cyclase. As Mel1c possesses an exceptionally large C-terminal tail, we further asked if this structural feature prevented productive coupling to Ga 16 . Eleven chimeric melatonin or mutant receptors were constructed by swapping all or part of the C-terminal tail between MT1, MT2 and Mel1c. The primary roles of melatonin, a hormone synthesized and secreted by the pineal gland, are to regulate behavioral and neuroendocrine processes, as well as circadian rhythms in mammals. High affinity melatonin receptors cloned from various organisms have been found to be homologs of one of three receptor subtypes, MT1, MT2 and Mel1c, which make up the G protein-coupled melatonin receptor family (Shiu et al. 1997). The mammalian MT1 and MT2 receptors are 60% identical to each other and they share 60% sequence homology with Xenopus Mel1c (Reppert et al. , 1995a. The receptor subtypes also have distinct expression patterns. Mammalian MT1 is expressed in the hypophyseal pars tuberalis and hypothalamic suprachiasmatic nuclei, regions of the brain that have been implicated as being important for reproduction and circadian rhythms, respectively . The mammalian MT2, on the other hand, is found in the retina (Reppert et al. 1995a). Furthermore, the three receptor subtypes may not necessarily occur in the same organisms. Initial studies indicate that only the MT1

G Protein Selectivity Is Regulated by Multiple Intracellular Regions of GPCRs

“…Truncation of the C-terminal tail of Mel 1c receptor, which couples to G i/o only, enabled the receptor to couple to G i/o and G 16 [37]. Crafting this C-terminal region of Mel 1c to the C-terminus of MT 1 or MT 2 , which couples to both G i/o and G 16 , disrupted the coupling to G 16 but not G i/o (table 4).…”

Role of melatonin on diabetes-related metabolic disorders

“…Furthermore, it was also shown that such a melatonininduced IP3 liberation was able to release Ca 2+ from intracellular stores [65] , a mechanism that is commonly accepted as a trigger for insulin secretion. Despite in vitro expressed melatonin receptors exhibiting differential abilities to stimulate phospholipase C (PLC) through Gq proteins [66,67] and the MT1 receptor having been shown to couple with Gq proteins in an agonist-dependent and guanine nucleotide-sensitive manner in HEK293 cells [68] , the melatonindependent stimulation of PLC through Gq-coupled MT1 receptor can only be hypothesized in pancreatic β-cells. In this regard, it has been reported that stimulation of INS1 cells with melatonin provokes the release of IP3 [65,69] , and when Gi coupling is blocked by pertussis toxin, a stimulatory effect of melatonin is revealed [69] .…”

Identification of a Novel Site within G Protein α Subunits Important for Specificity of Receptor-G Protein Interaction

“…Likewise, the first intracellular loop (Strader et al, 1994(Strader et al, , 1995Ho et al, 2002) or the receptor's C terminal tail (Perroy et al, 2001;Lai et al, 2002) rarely determine coupling specificity. On the level of the G protein ␣ subunit, several different regions have been implicated in recognition of GPCRs and thus determine the specificity of receptor-G protein coupling: the extreme C terminus (Conklin et al, 1993;Kostenis et al, 1997a;Blahos et al, 1998;Havlickova et al, 2003;, the extreme N terminus (Kostenis et al, 1997b;, a region between the ␣4-and ␣5-helices (␣4 -␤6 loop) (Mazzoni and Hamm, 1996;Bae et al, 1997;Onrust et al, 1997;, and a region within the loop linking the N-terminal ␣-helix to the ␤1-strand of the ras-like domain (Blahos et al, 2001).…”