Institute of Applied Biotechnology, Life Science and Technology School, Lingnan Normal University, Zhanjiang 524048, Guangdong, China; Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849, United States.
Institute of Applied Biotechnology, Life Science and Technology School, Lingnan Normal University, Zhanjiang 524048, Guangdong, China.
Gen Comp Endocrinol. 2019 Dec 1;284:113234. doi: 10.1016/j.ygcen.2019.113234. Epub 2019 Aug 6.
Melanocortin-4 receptor (MC4R) and melanocortin receptor accessory protein 2 (MRAP2) play important roles in the melanocortin system, and interaction of MC4R and MRAP2 is suggested to play pivotal role in energy balance of vertebrates. Orange-spotted grouper (Epinephelus coioides) is a widely cultured marine fish with high economic value in Asia. To explore potential interaction between grouper MC4R and MRAP2, herein we cloned grouper mc4r and mrap2. Grouper mc4r consisted of a 981 bp ORF encoding a putative protein of 327 amino acids, while the grouper mrap2 consisted of a 696 bp ORF encoding a putative protein of 232 amino acids. Sequence and phylogenetic analysis revealed that the grouper MC4R and MRAP2 were highly homologous at amino acid levels to several teleost MC4Rs and MRAP2s, respectively. qRT-PCR results showed that both mc4r and mrap2 were expressed primarily in the central nervous system. In the periphery, these genes were expressed more widely in male fish. The cloned grouper MC4R was functional, exhibiting high constitutive activity in cAMP pathway, capable of binding to three peptide agonists and increasing intracellular cAMP production dose-dependently. MRAP2 significantly decreased basal and agonist-stimulated cAMP signaling. MRAP2 also increased basal ERK1/2 activation but decreased ligand-induced stimulation when expressed at high levels. These data will facilitate future investigation of these molecules in regulating diverse physiological processes in orange-spotted grouper.
黑皮质素-4 受体 (MC4R) 和黑皮质素受体辅助蛋白 2 (MRAP2) 在黑皮质素系统中发挥重要作用，MC4R 和 MRAP2 的相互作用被认为在脊椎动物的能量平衡中发挥关键作用。橙点石斑鱼 (Epinephelus coioides) 是一种广泛养殖的海水鱼，在亚洲具有很高的经济价值。为了探索石斑鱼 MC4R 和 MRAP2 之间的潜在相互作用，本研究克隆了石斑鱼 mc4r 和 mrap2。石斑鱼 mc4r 由一个 981bp 的 ORF 组成，编码一个 327 个氨基酸的假定蛋白，而石斑鱼 mrap2 由一个 696bp 的 ORF 编码一个 232 个氨基酸的假定蛋白。序列和系统发育分析表明，石斑鱼 MC4R 和 MRAP2 在氨基酸水平上与几种硬骨鱼的 MC4R 和 MRAP2 高度同源。qRT-PCR 结果显示，mc4r 和 mrap2 主要在中枢神经系统中表达。在外周组织中，这些基因在雄性鱼类中表达更为广泛。克隆的石斑鱼 MC4R 具有功能，在 cAMP 途径中表现出高的组成型活性，能够结合三种肽激动剂并呈剂量依赖性地增加细胞内 cAMP 的产生。MRAP2 显著降低了基础和激动剂刺激的 cAMP 信号。当高表达时，MRAP2 还增加了基础 ERK1/2 的激活，但降低了配体诱导的刺激。这些数据将有助于未来研究这些分子在调控橙点石斑鱼多种生理过程中的作用。
