Abstract
Melanin-concentrating hormone (MCH) is a cyclic neuropeptide, with a major role in stimulation of feeding behavior in mammals. MCH signals in the brain occur via two seven-transmembrane G protein-coupled receptors, namely MCH1 (SLC-1, MCH1, MCH-R1, or MCH-1R) and MCH2 (SLT, MCH2, MCH-R2, or MCH-2R). In this study, we demonstrate that the pro-MCH131–165 peptide neuropeptide-glutamic acid-isoleucine (NEI)-MCH is more potent than MCH in stimulating feeding in the rat. Using rat MCH1-expressed human embryonic kidney 293 cells, we show that NEI-MCH exhibits 5-fold less affinity in a binding assay and 2-fold less potency in a cAMP assay than MCH. A similar 7- to 8-fold shift in potency was observed in a Ca2+i assay using rat MCH1 or human MCH2-transfected Chinese hamster ovary cell models. This demonstrates that NEI-MCH is not a better agonist than MCH at either of the MCH receptors. Then, we compared the proteolysis resistance of MCH and NEI-MCH to rat brain membrane homogenates and purified proteases. Kinetics of peptide degradation using brain extracts indicated at1/2 of 34.8 min for MCH and 78.5 min for NEI-MCH with a specific pattern of cleavage of MCH but not NEI-MCH by exo- and endo-proteases. Furthermore, MCH was found highly susceptible to degradation by aminopeptidase M and endopeptidase 24.11, whereas NEI-MCH was fully resistant to proteolysis by these enzymes. Therefore, our results strongly suggest that reduced susceptibility to proteases of NEI-MCH compared with MCH account for its enhanced activity in feeding behavior. NEI-MCH represents therefore the first MCH natural functional “superagonist” so far described.
Footnotes
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This work was supported in part by grants from the Institut de Recherche Servier, the Institut Danone, and the Centre National de la Recherche Scientifique to L.M.-F., T.S., C.R.-J., and J.L.N. L.M-F. and T.S. were recipients of “Nestle” and “Conventions Industrielles de Formation par la Recherche” (Association Nationale de la Recherche Technique, Institut de Recherche Servier, Centre National de la Recherche Scientifique) fellowships, respectively.
- Abbreviations:
- MCH
- melanin-concentrating hormone
- NEI
- neuropeptide-glutamic acid-isoleucine
- IR
- immunoreactive
- CSF
- cerebrospinal fluid
- BSA
- bovine serum albumin
- RP
- reverse phase
- HPLC
- high-performance liquid chromatography
- CHO
- Chinese hamster ovary
- PMSF
- phenylmethylsulfonyl fluoride
- MALDI-TOF
- matrix-assisted laser desorption ionization/time of flight
- MS
- mass spectroscopy
- Rt
- retention time
- [125I]S36057
- (S-S cyclic)(3-iodo)Tyr-8-amino-3,6-dioxyoctanoyl-Arg-Cys-Met-Leu-Gly-Arg-Val-Phe-Arg-Pro-Cys-Trp
- Received October 29, 2001.
- Accepted April 22, 2002.
- The American Society for Pharmacology and Experimental Therapeutics
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