Agouti protein, mahogunin, and attractin in pheomelanogenesis and melanoblast-like alteration of melanocytes: a cAMP-independent pathway

Xue, B. Z., W. O. Wilkison, R. L. Mynatt, N. Moustaid, M. Goldman, and M. B. Zemel. The agouti gene product stimulates pancreatic β-cell Ca2+ signaling and insulin release. Physiol. Genomics 1: 11-19, 1999.—Ubiquitous expression of the mouse agouti gene results in obesity and hyperinsulinemia. Human agouti is expressed in adipose tissue, and we found recombinant agouti protein to stimulate lipogenesis in adipocytes in a Ca2+-dependent fashion. However, adipocyte-specific agouti transgenic mice only became obese in the presence of hyperinsulinemia. Because intracellular Ca2+ concentration ([Ca2+]i) is a primary signal for insulin release, and we have shown agouti protein to increase [Ca2+]i in several cell types, we examined the effects of agouti on [Ca2+]i and insulin release. We demonstrated the expression of agouti in human pancreas and generated recombinant agouti to study its effects on Ca2+ signaling and insulin release. Agouti (100 nM) stimulated Ca2+ influx, [Ca2+]i increase, and a marked stimulation of insulin release in two β-cell lines (RIN-5F and HIT-T15; P < 0.05). Agouti exerted comparable effects in isolated human pancreatic islets and β-cells, with a 5-fold increase in Ca2+ influx ( P < 0.001) and a 2.2-fold increase in insulin release ( P < 0.01). These data suggest a potential role for agouti in the development of hyperinsulinemia in humans.

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Modulation of melanogenic protein expression during the switch from eu- to pheomelanogenesis

Journal of Cell Science ◽

10.1242/jcs.108.6.2301 ◽

1995 ◽

Vol 108 (6) ◽

pp. 2301-2309 ◽

Cited By ~ 2

Author(s):

T. Kobayashi ◽

W.D. Vieira ◽

B. Potterf ◽

C. Sakai ◽

G. Imokawa ◽

...

Keyword(s):

Matrix Protein ◽

Acid Oxidase ◽

Newborn Mice ◽

Dopachrome Tautomerase ◽

Melanocyte Stimulating Hormone ◽

Agouti Protein ◽

Enzymatic Function ◽

Lethal Yellow ◽

Related Proteins ◽

Catalytic Functions

Mammalian melanocytes can produce two basic types of melanin, eumelanin and pheomelanin, within discrete organelles termed melanosomes. The physiological signals that regulate this switch are extrinsic to the melanocyte, and include alpha-melanocyte stimulating hormone and the agouti protein. Tyrosinase, encoded at the albino locus, is the enzyme essential for the synthesis of both types of melanin, but other tyrosinase-related proteins (e.g. TRP1 encoded at the brown locus and TRP2 encoded at the slaty locus) regulate eumelanogenesis catalytically at steps distal to tyrosinase (as 5,6-dihydroxyindole-2-carboxylic acid oxidase and DOPAchrome tautomerase, respectively). The silver protein is another melanosomal protein, and although it has some limited homology to the tyrosinase-related proteins, it does not have any known enzymatic function and probably serves as a structural matrix protein. The role of each of those melanosomal proteins in pheomelanogenesis, however, is still unclear. In this study, we have compared the expression and catalytic functions of those proteins in pheomelanic and eumelanic hair bulb melanocytes. There was no detectable expression of TRP1 or TRP2, or either of their enzymatic activities, in hair bulbs of lethal yellow (Ay/a) newborn mice, and tyrosinase activity was present at a reduced level compared to that found in hair bulbs of black (a/a) newborn mice. Similar results were observed in regenerating hair bulbs of adult lethal yellow mice and in hair bulbs of 5- to 7-day-old agouti mice (A/A), an age where pheomelanin is produced predominantly. Expression of the silver protein was similarly not observed in hair bulbs of the pheomelanic mice.(ABSTRACT TRUNCATED AT 250 WORDS)

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Synergism Between Agouti Protein Fragment 91–131 and Melanin Concentrating Hormone

Peptides: The Wave of the Future ◽

10.1007/978-94-010-0464-0_332 ◽

2001 ◽

pp. 712-713

Author(s):

Alex N. Eberle ◽

Joszef Bódi ◽

György Orosz ◽

Helga Süli-Vargha ◽

Verena Jäggin ◽

...

Keyword(s):

Protein Fragment ◽

Agouti Protein ◽

Melanin Concentrating Hormone

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Opposite Orientations of an Inverted Duplication and Allelic Variation at the Mouse agouti Locus

Genetics ◽

10.1093/genetics/144.1.265 ◽

1996 ◽

Vol 144 (1) ◽

pp. 265-277

Author(s):

Yanru Chen ◽

David M J Duhl ◽

Gregory S Barsh

Keyword(s):

Homologous Recombination ◽

Allelic Variation ◽

Inbred Strains ◽

Mrna Isoforms ◽

Pigment Synthesis ◽

Agouti Locus ◽

Agouti Protein ◽

Hybridization Probes ◽

Inverted Duplication ◽

Untranslated Exon

Abstract The mouse agouti protein is a paracrine signaling molecule that causes yellow pigment synthesis. A pale ventral coloration distinguishes the light-bellied agouti (AW) from the agouti (A) allele, and is caused by expression of ventral-specific mRNA isoforms with a unique 5′ untranslated exon. Molecular cloning demonstrates this ventral-specific exon lies within a 3.1-kb element that is duplicated in the opposite orientation 15-kb upstream to produce an interrupted palindrome and that similarity between the duplicated elements has been maintained by gene conversion. Orientation of the palindrome is reversed in A compared to AW, which suggests that mutation from one allele to the other is caused by intrachromosomal homologous recombination mediated by sequences within the duplicated elements. Analysis of 15 inbred strains of laboratory and wild-derived mice with Southern hybridization probes and closely linked microsatellite markers suggests six haplotype groups: one typical for most strains that carry AW (129/SvJ, LP/J, CE/J, CAST/Ei), one typical for most strains that carry A (Balb/cJ, CBA/J, FVB/N, PERA/Rk, RBB/Dn); and four that are atypical (MOLC/Rk, MOLG/Dn, PERA/Ei, PERC/Ei, SPRET/Ei, RBA/Dn). Our results suggest a model for molecular evolution of the agouti locus in which homologous recombination can produce a reversible switch in allelic identity.

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Ectopic Agouti protein overexpression increases stimulated corticosterone production without effect on adenylate cyclase activity in mouse adrenal cells

Acta Endocrinologica ◽

10.1530/eje.0.1510613 ◽

2004 ◽

pp. 613-618 ◽

Cited By ~ 3

Author(s):

AY Shevchenko ◽

NM Bazhan ◽

EN Makarova ◽

TV Yakovleva ◽

NR Karkaeva

Keyword(s):

Adenylate Cyclase ◽

Protein Overexpression ◽

Dibutyryl Camp ◽

Steroidogenic Enzymes ◽

Steroidogenic Enzyme ◽

Adrenal Cells ◽

Agouti Protein ◽

Corticosterone Release ◽

Agouti Related Protein ◽

Enzyme Reactivity

OBJECTIVE: The antagonism of Agouti protein (AP) and Agouti-related protein on melanocortin receptors suggests an inhibitory role in the regulation of steroidogenesis. However, we have previously demonstrated that ectopic AP overexpression increased restraint-induced corticosterone release and adrenal reactivity to ACTH in mice. A high steroidogenic response to ACTH may be a consequence of a stimulatory AP action on the adenylate cyclase (AC) and/or intracellular steroidogenic enzymes. The aim of the present study was to estimate the effect of ectopic AP overexpression on the activity of AC and steroidogenic intracellular enzymes. METHODS: ACTH and forskolin were used for AC stimulation, and dibutyryl cAMP and progesterone were used for stimulation of intracellular steroidogenic enzymes in isolated adrenal cells in male C57Bl/6J mice of two Agouti genotypes: A(y)/a (ectopic AP overexpression) and a/a (absence of AP in all tissues). RESULTS: ACTH and forskolin increased cAMP accumulation to the same extent in both A(y)/a and a/a mouse adrenal cells (P<0.001; ANOVA), but resulted in higher corticosterone production in A(y)/a mice (P<0.001 for ACTH and P<0.01 for forskolin; ANOVA). Dibutyryl cAMP- and progesterone-induced corticosterone production was higher in A(y)/a mice than in a/a mice (P<0.001 for dibutyryl cAMP and P<0.01 for progesterone; ANOVA). CONCLUSIONS: Ectopic AP overexpression increased stimulated corticosterone production and intracellular steroidogenic enzyme reactivity to cAMP without an effect on AC activity.

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Leptin responsiveness in mice that ectopically express agouti protein

Physiology & Behavior ◽

10.1016/s0031-9384(01)00653-9 ◽

2002 ◽

Vol 75 (1-2) ◽

pp. 159-167 ◽

Cited By ~ 11

Author(s):

Ruth B.S Harris ◽

Tiffany D Mitchell ◽

Randall L Mynatt

Keyword(s):

Agouti Protein

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Dose–response effects of ectopic agouti protein on iron overload and age-associated aspects of the Avy/a obese mouse phenome

Peptides ◽

10.1016/j.peptides.2004.12.033 ◽

2005 ◽

Vol 26 (10) ◽

pp. 1697-1711 ◽

Cited By ~ 3

Author(s):

George L. Wolff ◽

Paul Whittaker

Keyword(s):

Iron Overload ◽

Dose Response ◽

Obese Mouse ◽

Agouti Protein

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Melanocortin receptor-mediated mobilization of intracellular free calcium in HEK293 cells

Physiological Genomics ◽

10.1152/physiolgenomics.2001.5.1.11 ◽

2001 ◽

Vol 5 (1) ◽

pp. 11-19 ◽

Cited By ~ 69

Author(s):

KATHLEEN G. MOUNTJOY ◽

PHILIP L. KONG ◽

JOHN A. TAYLOR ◽

DERRIL H. WILLARD ◽

WILLIAM O. WILKISON

Keyword(s):

Intracellular Free Calcium ◽

Melanocortin Receptor ◽

Hek293 Cells ◽

Melanocortin Receptors ◽

Calcium Signal ◽

Free Calcium ◽

Agouti Protein ◽

Transient Rise ◽

Physiological Relevance ◽

Free Calcium Concentration

Mouse melanocortin receptors, MC1-R, MC3-R, MC4-R, and MC5-R, when expressed in HEK293 cells and stimulated with either α-melanocyte-stimulating hormone (α-MSH) or desacetyl-α-MSH, mediate increases in intracellular free calcium concentration ([Ca2+]i) with EC50 values between 0.3 and 4.3 nM. The increase in [Ca2+]i is cholera toxin sensitive and pertussis toxin insensitive. The mechanism involves calcium mobilization from intracellular stores without a transient rise in inositol trisphosphate. Mouse agouti protein (55 nM) is a competitive antagonist of α-MSH (6-fold) and desacetyl-α-MSH (8-fold), coupling the mMC1-R to increased [Ca2+]i. Agouti protein (55 nM) significantly increased the EC50 for α-MSH (3-fold), and 550 nM agouti protein significantly increased the EC50 for desacetyl-α-MSH (4-fold), coupling the mMC4-R to a rise in [Ca2+]i. However, agouti protein antagonism of the MC4-R may not be competitive since there was a trend for the maximum response to also increase. There was no significant antagonism of the MC3-R and MC5-R by agouti protein (55 nM). Understanding the physiological relevance of the transduction of a calcium signal by melanocortin peptides may be important for future development of therapeutic targeting of the melanocortin receptors.

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A transgenic mouse assay for agouti protein activity.

Genetics ◽

10.1093/genetics/140.1.267 ◽

1995 ◽

Vol 140 (1) ◽

pp. 267-274

Author(s):

W L Perry ◽

C M Hustad ◽

D A Swing ◽

N A Jenkins ◽

N G Copeland

Keyword(s):

Transgenic Mice ◽

Signal Sequence ◽

Middle Part ◽

Yellow Pigment ◽

Black Hair ◽

In Vitro Mutagenesis ◽

Protein Activity ◽

Functional Elements ◽

Agouti Protein

Abstract The mouse agouti gene encodes an 131 amino acid paracrine signaling molecule that instructs hair follicle melanocytes to switch from making black to yellow pigment. Expression of agouti during the middle part of the hair growth cycle in wild-type mice produces a yellow band on an otherwise black hair. The ubiquitous unregulated expression of agouti in mice carrying dominant yellow alleles is associated with pleiotropic effects including increased yellow pigment in the coat, obesity, diabetes and increased tumor susceptibility. Agouti shows no significant homology to known genes, and the molecular analysis of agouti alleles has shed little new light on the important functional elements of the agouti protein. In this paper, we show that agouti expression driven by the human beta-ACTIN promoter produces obese yellow transgenic mice and that this can be used as an assay for agouti activity. We used this assay to evaluate a point mutation associated with the a16H allele within the region encoding agouti's putative signal sequence and our results suggest that this mutation is sufficient to cause the a16H phenotype. Thus, in vitro mutagenesis followed by the generation of transgenic mice should allow us to identify important functional elements of the agouti protein.

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