Molecular cloning and expression of two melanin-concentrating hormone receptors in goldfish

• Published in: Peptides (New York, N.Y. : 1980) Vol. 30; no. 11; pp. 1990 - 1996
• Main Authors: Mizusawa, Kanta, Saito, Yumiko, Wang, Zhiwei, Kobayashi, Yuki, Matsuda, Kouhei, Takahashi, Akiyoshi
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.11.2009
• Subjects: Actins - metabolism; Amino Acid Sequence; Animals; Brain - metabolism; Calcium - metabolism; Calcium influx; Carassius auratus; Cell Line; Freshwater; G-protein-coupled receptor; Gene Expression Regulation; Goldfish; Goldfish - genetics; Goldfish - metabolism; Humans; Melanin-concentrating hormone; Molecular Sequence Data; Pertussis toxin; Phylogeny; Receptors, G-Protein-Coupled - genetics; Receptors, G-Protein-Coupled - metabolism; Receptors, Pituitary Hormone - chemistry; Receptors, Pituitary Hormone - genetics; Receptors, Pituitary Hormone - metabolism; Reverse Transcriptase Polymerase Chain Reaction; Salmonidae; Sequence Alignment; Sequence Homology, Amino Acid; Skin - metabolism; Teleostei; Goldfish; Melanin-concentrating hormone; Pertussis toxin; Calcium influx; G-protein-coupled receptor
• ISSN: 0196-9781; 1873-5169
• DOI: 10.1016/j.peptides.2009.04.010

Melanin-concentrating hormone (MCH) is a neurohypophysial hormone and induces melanin aggregation in the skin in teleosts. MCH also has multiple roles in the central regulation of food intake in teleosts and mammals. MCH receptors (MCH-R) are among type I G-protein-coupled receptors. Here, we cloned two MCH receptors from goldfish, Carassius auratus. The amino acid sequence of goldfish MCH-R1 had 57–88% homology with fish MCH-R1 and 49–50% homology with mammalian MCH-R1, while the amino acid sequence of goldfish MCH-R2 had 72–92% homology with fish MCH-R2 and 32% homology with human MCH-R2. Phylogenetic analysis showed that these two MCH-Rs are orthologous to the respective mammalian MCH-Rs. The common amino acid residues for ligand binding, signal transduction, and receptor conformation were well conserved in these receptors, although some intracellular basic-amino-acid-rich domains, which have been shown to exist in human MCH-R1 and MCH-R2, were absent in goldfish MCH-R2. When stably expressed in HEK293 cells, both goldfish MCH-R1 and MCH-R2 displayed a strong, dose-dependent, transient elevation of intracellular calcium in response to salmon MCH (EC 50 = 0.8 nM and 31.8 nM, respectively). In contrast to goldfish MCH-R2, goldfish MCH-R1 signaling is not sensitive to pertussis toxin, suggesting an exclusive Gαq coupling of goldfish MCH-R1 in the mammalian cell-based assay. Reverse transcriptase PCR revealed that both MCH-R1 and MCH-R2 mRNA are distributed in various tissues in goldfish. The various tissues including the brain and skin express both MCH-R1 and MCH-R2. These results suggest that these functional receptors mediate multiple effects of MCH in goldfish.