Agouti regulation of intracellular calcium: role of melanocortin receptors.
Several dominant mutations at the murine agouti locus cause a syndrome of marked obesity and insulin resistance. We have recently reported that intracellular free Ca2+ concentration ([Ca2+]i) is elevated in viable yellow mice. Because [Ca2+]i has a key role in the pathogenesis of insulin resistance, obesity, and hypertension, the role of the purified agouti gene product in regulating [Ca2+]i was evaluated in a number of cell types. Purified murine agouti induced slow, sustained increases in [Ca2+]i in A7r5 vascular smooth muscle cells and 3T3-L1 adipocytes in a dose-dependent fashion. In L6 skeletal myocytes, agouti stimulated an increase in [Ca2+]i with an apparent concentration eliciting 50% of the maximal response (EC50) of 62 nM. This response was substantially inhibited by Ca2+ entry blockade with nitrendipine. To determine whether melanocortin receptors play a role in agouti regulation of [Ca2+]i, we examined the effect of melanocortin peptides and agouti in cells stably transfected with human melanocortin receptors. Human embryonic kidney cells (HEK-293 cells) transfected with either the human melanocortin 1 receptor (MC1R) or melanocortin 3 receptor responded to human agouti with slow, sustained increases in [Ca2+]i, whereas nontransfected HEK-293 cells with no melanocortin receptors did not respond to agouti. Dose-response curves in the MC1R line showed that agouti had an EC50 of 18 nM, which is comparable to that for agouti antagonism of (125)I-Nle,D-Phe-alpha-melanocyte-stimulating hormone binding in the same cell line. This direct effect of agouti on stimulating increases in [Ca2+]i suggests a potential mechanism for agouti-induced insulin resistance.
Kim
J H
JH
Department of Nutrition and Physiology Program, University of Tennessee, Knoxville 37996-1900, USA.
Kiefer
L L
LL
Woychik
R P
RP
Wilkison
W O
WO
Truesdale
A
A
Ittoop
O
O
Willard
D
D
Nichols
J
J
Zemel
M B
MB
D017667:Adipocytes / Q000378:metabolism; D054366:Agouti Signaling Protein; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D009074:Melanocyte-Stimulating Hormones / Q000494:pharmacology; D051379:Mice; D008817:Mice, Mutant Strains; D018482:Muscle, Skeletal / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D011506:Proteins / Q000494:pharmacology*; D044104:Receptor, Melanocortin, Type 3; D018043:Receptors, Corticotropin / Q000502:physiology*; D044101:Receptors, Melanocortin; D011994:Recombinant Proteins; D014162:Transfection
D054366:Agouti Signaling Protein; D036341:Intercellular Signaling Peptides and Proteins; C470053:Mc3r protein, mouse; D011506:Proteins; D044104:Receptor, Melanocortin, Type 3; D018043:Receptors, Corticotropin; D044101:Receptors, Melanocortin; D011994:Recombinant Proteins; D009074:Melanocyte-Stimulating Hormones; D002118:Calcium