Melanocortin-4 receptor in swamp eel (Monopterus albus): Cloning, tissue distribution, and pharmacology

Yi, Ti-Lin; Yang, Likun; Ruan, Guoliang; Yang, Dan; Tao, Ya-Xiong

doi:10.1016/j.gene.2018.07.056

Cited by 26 publications

(33 citation statements)

References 60 publications

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“…In the present study, we demonstrated that cloned culter mc4r was predicted to encode a protein of 326 amino acids with similar structural characteristics as MC4Rs of other species (Figure 1A and Supplementary Figure 2), including other teleost MC4Rs (27)(28)(29)(30)(31)37). Phylogenetic analysis revealed that caMC4R clustered with teleost MC4Rs (Figure 1B).…”

Section: Discussionmentioning

confidence: 56%

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Regulation of Melanocortin-4 Receptor Pharmacology by Two Isoforms of Melanocortin Receptor Accessory Protein 2 in Topmouth Culter (Culter alburnus)

Tao

Huang

et al. 2020

Front. Endocrinol.

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Melanocortin-4 receptor (MC4R) plays important roles in regulation of multiple physiological processes, and interaction of MC4R and melanocortin receptor accessory protein 2 (MRAP2) is suggested to play pivotal role in energy balance of vertebrates. Topmouth culter (Culter alburnus) is an economically important freshwater fish in China. Herein we cloned culter mc4r, mrap2a, and mrap2b. Culter mc4r consisted of a 981 bp open reading frame encoding a protein of 326 amino acids. qRT-PCR revealed that mc4r, mrap2a, and mrap2b were primarily expressed in the central nervous system. In the periphery, mc4r and mrap2b were expressed more widely in the male, while mrap2a was expressed more widely in the female. Culter MC4R could bind to four peptide agonists and increase intracellular cAMP production dose dependently. Culter MC4R was constitutively active in both cAMP and ERK1/2 pathways, which was differentially regulated by culter MRAP2a and MRAP2b. Culter MRAP2a significantly increased B max and decreased agonist-stimulated cAMP, while MRAP2b increased cell surface and total expression but did not affect B max and agonist-stimulated cAMP. These results will aid the investigation of the potential physiological processes that MC4R might be involved in topmouth culter.

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Section: Discussionmentioning

confidence: 56%

“…MC4R has also been characterized in teleosts (21,(24)(25)(26)(27)(28)(29)(30)(31)(32), and shown to act as a regulator in energy balance, sexual behavior, and reproduction (33)(34)(35)(36)(37). Non-mammalian MRAPs are different from those of mammals.…”

Section: Introductionmentioning

confidence: 99%

Regulation of Melanocortin-4 Receptor Pharmacology by Two Isoforms of Melanocortin Receptor Accessory Protein 2 in Topmouth Culter (Culter alburnus)

Tao

Huang

et al. 2020

Front. Endocrinol.

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“…It is also possible that D146 is the residue or one of the residues critical for contacting Ipsen 5i and ML00253764. In fish MC4Rs, Ipsen 5i also works as an allosteric agonist [ 41 , 42 ]. In one previous study, an L133M mutation of MC4R also converts an antagonistic effect of SHU9199 to an agonistic effect [ 43 ].…”

Section: Discussionmentioning

confidence: 99%

Alanine Scanning Mutagenesis of the DRYxxI Motif and Intracellular Loop 2 of Human Melanocortin-4 Receptor

Yang

Tao

2020

IJMS

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The melanocortin-4 receptor (MC4R) is a member of the G-protein-coupled receptor (GPCR) superfamily, which has been extensively studied in obesity pathogenesis due to its critical role in regulating energy homeostasis. Both the Gs-cAMP and ERK1/2 cascades are known as important intracellular signaling pathways initiated by the MC4R. The DRYxxI motif at the end of transmembrane domain 3 and the intracellular loop 2 (ICL2) are thought to be crucial for receptor function in several GPCRs. To study the functions of this domain in MC4R, we performed alanine-scanning mutagenesis on seventeen residues. We showed that one residue was critical for receptor cell surface expression. Eight residues were important for ligand binding. Mutations of three residues impaired Gs-cAMP signaling without changing the binding properties. Investigation on constitutive activities of all the mutants in the cAMP pathway revealed that six residues were involved in constraining the receptor in inactive states and five residues were important for receptor activation in the absence of an agonist. In addition, mutations of four residues impaired the ligand-stimulated ERK1/2 signaling pathway without affecting the binding properties. We also showed that some mutants were biased to the Gs-cAMP or ERK1/2 signaling pathway. In summary, we demonstrated that the DRYxxI motif and ICL2 were important for MC4R function.

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“…Cycling conditions were as follows: 95 °C for 3 min, followed by 40 cycles of 95 °C for 10 s, 55.0 °C for 20 s, 72.0 °C for 20 s, and plate reading at 75 °C for 5 s. qPCR specificity was assessed using melt curves (65–95 °C, with 0.5 °C increments) and electrophoresis. Previous studies in this species mostly relied on β‐actin as the reference gene , but we also tested the expression of another commonly used reference gene, GAPDH , in different tissues. Stability of Ct values of these two genes was analyzed using the comparative cycle threshold method implemented in the GeNorm software .…”

Section: Methodsmentioning

confidence: 99%

“…cDNA library was synthesized using RevertAid TM First Strand cDNA Synthesis Kit with DNase I (#K1621 and #EN0521, respectively, both Thermo Scientific) following the manufacturer's protocol, and cDNA stored at À80°C. As we could identify only one kiss/gpr54 gene using the methodology described in the first section, to further confirm that there are no other expressed kiss/gpr54 paralogues present in the transcriptome of M. albus (most importantly, to corroborate that kiss1 and gpr54-1 are absent), we designed degenerate primers for kiss1, kiss2, and gpr54 genes ( [25,26], but we also tested the expression of Inter is intersex, H is heart, L is liver, S is spleen, K is kidney, B is brain, G is gonad, M is muscle, P is pituitary gland, and I is intestine. Discarded samples are highlighted with a red X sign above.…”

Section: Rna Cdna and Racementioning

confidence: 99%

Expression patterns of kiss2 and gpr54‐2 in Monopterus albus suggest these genes may play a role in sex reversal in fish

Pei²,

Yang

2019

FEBS Open Bio

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Due to its exceptionally small genome size and protogynous hermaphroditism, Monopterus albus has been proposed as a model for vertebrate sexual development. The Kiss/GPR54 system is a central regulator of sexual development in most vertebrates, but its role in sex reversal remains hypothetical. In contrast to mammals, fishes often possess more than one copy of the kiss and gpr54 genes. Our objectives were to identify all kiss/gpr54 genes in the genome of M. albus and to assess their involvement in sex reversal via their expression patterns (qPCR) in females, males, and intersex specimens. We identified only two genes: kiss2 and gpr54‐2. kiss2 expression was extremely high in the gonads of males, intermediate in females, and low in intersex; and reduced in all tissues of intersex. gpr54 expression was also extremely high in the gonads of males, high in intersex, but low in females. gpr54 expression in brain was high in all three sexes. In conclusion, (a) kiss1 has been functionally replaced in M. albus; (b) the functions of gpr54‐2 in brain are not sex‐specific; (c) kiss2 appears to undergo a ‘reset’ in the expression during the sex change; and (d) sex‐specific expression patterns in the gonads indicate that these two genes may play a role in sex reversal in fish.

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Melanocortin-4 receptor in swamp eel (Monopterus albus): Cloning, tissue distribution, and pharmacology

Cited by 26 publications

References 60 publications

Regulation of Melanocortin-4 Receptor Pharmacology by Two Isoforms of Melanocortin Receptor Accessory Protein 2 in Topmouth Culter (Culter alburnus)

Regulation of Melanocortin-4 Receptor Pharmacology by Two Isoforms of Melanocortin Receptor Accessory Protein 2 in Topmouth Culter (Culter alburnus)

Alanine Scanning Mutagenesis of the DRYxxI Motif and Intracellular Loop 2 of Human Melanocortin-4 Receptor

Expression patterns of kiss2 and gpr54‐2 in Monopterus albus suggest these genes may play a role in sex reversal in fish

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