title: Pharmacologic analyses of four chicken melanocortin-4 receptor mutations.
abstract: The melanocortin-4 receptor (MC4R) is a critical regulator of mammalian food intake and energy expenditure, with receptor activation resulting in decreased food intake and increased energy expenditure. Recently, studies on role of MC4R in regulation of food intake have been extended to other species, such as chicken. Functional study of mutant MC4Rs is important in proving the causal link between MC4R mutation and production traits. Herein, we cloned chicken MC4R (cMC4R) complementary DNA and generated 4 mutant cMC4Rs (Q18H, G21R, S76L, and L299P) by site-directed mutagenesis and measured their expression by flow cytometry. Pharmacologic characteristics were analyzed with binding and signaling assays using 3 agonists. We showed that G21R had decreased cell surface and total expression (P < 0.05), whereas the other 3 mutants had similar total and cell surface expression levels as wild-type cMC4R. The 4 mutants had either decreased (Q18H, G21R, S76L; P < 0.05) or no (L299P) binding to radiolabeled [Nle(4), D-Phe(7)]-α-melanocyte-stimulating hormone (MSH). In signaling assays, Q18H was constitutively active. Q18H, G21R, and S76L had decreased responses to α-MSH stimulation (P < 0.05). L299P had decreased basal and ligand-stimulated signaling (P < 0.01). Nle(4), D-Phe(7)-MSH was the most potent agonist for cMC4R and therefore would be better suited for further in vivo studies. We conclude that the cloned cMC4R was a functional receptor and provided detailed functional data for these mutations, contributing to a better understanding of cMC4R variants associated with production traits.
authors.affiliation: College of Veterinary Medicine, Yangzhou University, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu 225009, People's Republic of China; Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849, USA. Electronic address: zqwang@yzu.edu.cn.
authors.forename: Z-Q
authors.initials: ZQ
authors.lastname: Wang
authors.affiliation: Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849, USA.
authors.forename: J-S
authors.initials: JS
authors.lastname: Huang
authors.affiliation: College of Veterinary Medicine, Yangzhou University, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu 225009, People's Republic of China.
authors.forename: J-H
authors.initials: JH
authors.lastname: Zhou
authors.affiliation: College of Veterinary Medicine, Yangzhou University, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu 225009, People's Republic of China.
authors.forename: L
authors.initials: L
authors.lastname: Shi
authors.affiliation: College of Veterinary Medicine, Yangzhou University, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu 225009, People's Republic of China.
authors.forename: X-F
authors.initials: XF
authors.lastname: Jiang
authors.affiliation: Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849, USA. Electronic address: taoyaxi@auburn.edu.
authors.forename: Y-X
authors.initials: YX
authors.lastname: Tao
mesh_terms: D000818:Animals; D030161:Avian Proteins / Q000235:genetics* / Q000378:metabolism; D002645:Chickens; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000096:biosynthesis; D018076:DNA, Complementary / Q000235:genetics; D004435:Eating / Q000502:physiology; D004734:Energy Metabolism / Q000502:physiology; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D044105:Receptor, Melanocortin, Type 4 / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D000521:alpha-MSH / Q000494:pharmacology
chemical_list: D030161:Avian Proteins; D018076:DNA, Complementary; D044105:Receptor, Melanocortin, Type 4; D000521:alpha-MSH; D000242:Cyclic AMP
keywords: Chicken; Ligand binding; Melanocortin-4 receptor; Mutation; Signaling