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Data Brief. 2016 June; 7: 1670–1677.
Published online 2016 April 26. doi:  10.1016/j.dib.2016.04.050
PMCID: PMC4927774

Data for amino acid alignment of Japanese stingray melanocortin receptors with other gnathostome melanocortin receptor sequences, and the ligand selectivity of Japanese stingray melanocortin receptors

Abstract

This article contains structure and pharmacological characteristics of melanocortin receptors (MCRs) related to research published in “Characterization of melanocortin receptors from stingray Dasyatis akajei, a cartilaginous fish” (Takahashi et al., 2016) [1]. The amino acid sequences of the stingray, D. akajei, MC1R, MC2R, MC3R, MC4R, and MC5R were aligned with the corresponding melanocortin receptor sequences from the elephant shark, Callorhinchus milii, the dogfish, Squalus acanthias, the goldfish, Carassius auratus, and the mouse, Mus musculus. These alignments provide the basis for phylogenetic analysis of these gnathostome melanocortin receptor sequences. In addition, the Japanese stingray melanocortin receptors were separately expressed in Chinese Hamster Ovary cells, and stimulated with stingray ACTH, α-MSH, β-MSH, γ-MSH, δ-MSH, and β-endorphin. The dose response curves reveal the order of ligand selectivity for each stingray MCR.

Keywords: Adrenocorticotropic hormone (ACTH), Dasyatis akajei, Melanocortin receptor (MCR), Melanocyte-stimulating hormone (MSH), Stingray

Specifications Table

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Value of the data

  • • These data are valuable for researchers participated in endocrinology of primitive fish and evolution of melanocortin systems.
  • • These could be used as probes to explore orthologs in other cartilaginous fish such as skates, sharks and chimaeras.
  • • The data on ligand selectivity could be useful tools for structure–function relationship studies in endocrinology and pharmacology.

1. Data

Data provided in this article show amino acid sequence comparison of melanocortin receptors (MCRs) in vertebrates and ligand selectivity of stingray MC peptides on these receptors. The amino acids sequences of MC1R (Fig. 1), MC2R (Fig. 2), MC3R (Fig. 3), MC4R (Fig. 4), and MC5R (Fig. 5) of stingray (Squalus acanthias) which determined by us [1] were compared to corresponding sequences from two species of other cartilaginous fishes (i.e., Callorhinchus milii, elephant shark and S. acanthias, dogfish), a teleost (Carassius auratus, goldfish), and a mammal (Mus musculus, mouse). Data are also provided for ligand selectivity include effects of stingray Des-acetyl-α-MSH, β-MSH, γ-MSH, δ-MSH, ACTH(1-24) and β-endorphin on MC1R, MC3R, MC4R, and MC5R (Fig. 6, Fig. 8, Fig. 9, Fig. 10) and those of stingray Des-acetyl-α-MSH, ACTH(1-24), human ACTH(1-24) and NDP-MSH on stingray MC2R (Fig. 7).

Fig. 1
Amino acid sequence comparison of MC1R used for phylogenetic analysis. Species names are Dasyatis akajei for stingray, Callorhinchus milii for elephant shark, Carassius auratus for goldfish, and Mus musculus for mouse. Dot shows identical amino acid to ...
Fig. 2
Amino acid sequence comparison of MC2R used for phylogenetic analysis. Species names are Dasyatis akajei for stingray, Callorhinchus milii for elephant shark, Carassius auratus for goldfish, and Mus musculus for mouse. Dot shows identical amino acid to ...
Fig. 3
Amino acid sequence comparison of MC3R used for phylogenetic analysis. Species names are Dasyatis akajei for stingray, Callorhinchus milii for elephant shark, Squalus acanthias for dogfish, Carassius auratus for goldfish, and Mus musculus for mouse. Dot ...
Fig. 4
Amino acid sequence comparison of MC4R used for phylogenetic analysis. Species names are Dasyatis akajei for stingray, Squalus acanthias for dogfish, Carassius auratus for goldfish, and Mus musculus for mouse. Dot shows identical amino acid to stingray ...
Fig. 5
Amino acid sequence comparison of MC5R used for phylogenetic analysis. Species names are Dasyatis akajei for stingray, Squalus acanthias for dogfish, Carassius auratus for goldfish, and Mus musculus for mouse. Dot shows identical amino acid to stingray ...
Fig. 6
Ligand selectivity of stingray MC1R. (A) Functional activation of the stingray MC1R after stimulation with the following stingray melanocortins: Des-acetyl-α-MSH (Des-Ac-α-MSH), ACTH(1-24), or β-MSH. (B) Functional activation of ...
Fig. 7
Ligand selectivity of stingray MC2R. (A) Functional activation of the stingray MC2R after stimulation with stingray Des-acetyl-α-MSH (Des-Ac-α-MSH) or stingray ACTH(1-24). (B) Functional activation of stingray MC2R after stimulation with ...
Fig. 8
Ligand selectivity of stingray MC3R. (A) Functional activation of the stingray MC3R after stimulation with the following stingray melanocortins: Des-acetyl-α-MSH (Des-Ac-α-MSH), ACTH(1-24), or β-MSH. (B) Functional activation of ...
Fig. 9
Ligand selectivity of stingray MC4R. (A) Functional activation of the stingray MC4R after stimulation with the following stingray melanocortins: Des-acetyl-α-MSH (Des-Ac-α-MSH), ACTH(1-24), or β-MSH. (B) Functional activation of ...
Fig. 10
Ligand selectivity of stingray MC5R. (A) Functional activation of the stingray MC5R after stimulation with the following stingray melanocortins: Des-acetyl-α-MSH (Des-Ac-α-MSH), ACTH(1-24), or β-MSH. (B) Functional activation of ...

2. Experimental design, materials and methods

In order to align the amino acid sequences of the melanocortin receptors for the Japanese stingray, D. akajei, the dogfish, S. acanthias, the elephant shark, C. milii, the goldfish, C. auratus, and the mouse, M. musculus, it was essential to identify putative transmembrane domains in each receptor sequence. To this end, the program ‘‘MEMSAT3’’ (http://bioinf.cs.ucl.ac.uk/psipred/) was used. The amino acid sequences where then aligned using the program MEGA 6.0.

To functionally express and determine the ligand selectivity of the stingray (sr) MC1R, srMC2R, srMC3R, srMC4R, and srMC5R paralogs, the nucleotide sequences for the srmcrs were separately synthesized with a V-5 epitope tag at the N-terminal of the receptor, and inserted into a pcDNA3.1 expression vector (GenScript; Picataway, NJ, USA). Each srmcr cDNA was separately transiently transfected into Chinese Hamster Ovary (CHO) cells. The CHO cells were grown at 37 °C in a humidified 5% CO2 incubator in DMEM/F12 with 5% fetal calf serum. Each sr cDNA was co-expressed with a CRE/Luciferase reporter plasmid [2] using the Solution T Cell Line Nucleofector Kit (Amaxa Inc., Gaithersburg, MD, USA) and program U-23 [4]. The transiently transfected cells were seeded on a 96-well plate at a density of 1×10−5 cells/well. After 48 h in culture, the transfected cells were stimulated with either synthetic srACTH(1-24), srDes-acetyl-α-MSH, srβ-MSH, srγ-MSH, srδ-MSH, srβ-endorphin or hACTH(1-24), or NDP-MSH at concentrations ranging from 10−6 M to 10−12 M, in serum-free CHO media for four hours at 37 °C. At the end of the incubation period, 100 µl of Bright-Glo luciferase assay reagent (Promega Inc., Madison, WI, USA) was added to each well, and incubated for 5 min at room temperature. Luminescence was measured with a Bio-Tek Synergy HT plate reader (Bio Tek, Winooski, VT, USA), and the dose response curves were analyzed by using Kaleidagraph software (Synergy Software, Reading, PA, USA). All experimental treatments were performed in triplicate.

Acknowledgments

We thank Mr. Kosuke Arai, Kitasato University, and Dr. Yasuhisa Kobayashi, Dr. Naoaki Tsutsui and Mr. Kazuhiro Saito, Okayama University, for technical assistance. Partial support for this study was provided by the Long Research Fund (RMD) and National Science Foundation, U.S.A. Grant IOB 0516958 Supplement (RMD).

Footnotes

Appendix ASupplementary data associated with this article can be found in the online version at doi:10.1016/j.dib.2016.04.050.

Appendix A. Supplementary material

Supplementary material

References

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2. Chepurny O.G., Holz G.G. A novel cyclic adenosine monophosphate responsive luciferase reporter incorporating a nonpalindromic cyclic adenosine monophosphate response element provides optimal performance for use in G protein coupled receptor drug discovery efforts. J. Biomol. Screen. 2007;12:740–746. [PubMed]
3. Liang L., Sebag J.A., Eagelston L., Serasinghe M.N., Veo K., Reinick C., Angelson J., Hinkle P.M., Dores R.M. Functional expression of frog and rainbow trout melanocortin 2 receptors using heterologous MRAP1s. Gen. Comp. Endocrinol. 2011;174:5–14. [PubMed]
4. Reinick C.L., Liang L., Angleson J.K., Dores R.M. Identification of an MRAP-independent melanocortin-2 receptor: functional expression of the cartilaginous fish, Callorhinchus milii, melanocortin-2 receptor in CHO cells. Endocrinology. 2012;153:4757–4765. [PubMed]

Articles from Data in Brief are provided here courtesy of Elsevier