The interaction of MC3R and MC4R with MRAP2, ACTH, α-MSH and AgRP in chickens.
The interaction of MC3R and MC4R with MRAP2, ACTH, α-MSH and AgRP in chickens.
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DOI：
10.1530/joe-17-0131
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发表时间：
2017-05
期刊：
影响因子：
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通讯作者：
Jiannan Zhang (张剑南);Xin Li;Yawei Zhou;Lin Cui;Jing Li;Chenlei Wu;Y. Wan;Juan Li;Yajun Wang-Yaju
中科院分区：
文献类型：
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作者：
Jiannan Zhang (张剑南);Xin Li;Yawei Zhou;Lin Cui;Jing Li;Chenlei Wu;Y. Wan;Juan Li;Yajun Wang-Yaju
黑素皮质素-4（MC 4 R）和黑素皮质素-3（MC 3R）受体与阿黑皮素原（POMC）衍生肽（如α-MSH）、刺鼠相关蛋白（AgRP）和黑素皮质素-2受体辅助蛋白2（MRAP 2）的相互作用被认为在脊椎动物能量平衡中起关键作用。然而，关于它们在鸟类中相互作用的证据仍然很少。结果表明：（1）鸡（c-）MC 3R和cMC 4 R在中国仓鼠卵巢（CHO）细胞中的表达可被α-MSH和ACTH 1 -39等电位激活，并通过pGL 3-CRE-荧光素酶报告系统进行监测;（2）cMC 3R和cMC 4 R，当与cMRAP 2共表达时（或cMRAP，cMRAP 2同源物），显示对ACTH治疗的敏感性增加，因此可能作为ACTH偏好受体，免疫共沉淀法证实cMC 3R/cMC 4 R与cMRAP 2之间存在相互作用;（3）双荧光素酶报告基因检测表明cMC 3R和cMC 4 R在CHO细胞中表达时均表现出组成型活性，cMRAP 2在CHO细胞中表达时表现出组成型活性，（和cMRAP）可以调节它们的组成型活性;（4）AgRP抑制cMC 3R/cMC 4 R的组成性活性，并拮抗ACTH/α-MSH对cMC 4 R/cMC 3R的作用，表明AgRP作为两种受体的反向激动剂和拮抗剂起作用。这些结果以及实时荧光定量PCR检测到的鸡下丘脑中cMC 4 R、cMC 3R、cMRAP 2、cAgRP和cPOMC的共表达表明，在下丘脑中，α-MSH/ACTH、AgRP和MRAP 2可能在MC 4 R（/MC 3R）界面相互作用，控制能量平衡。此外，我们的数据提供了新的证据，MRAP 2（和MRAP）参与微调的组成活性和配体的敏感性和选择性的MC 3R和MC 4 R在脊椎动物。
The interaction of melanocortin-4 (MC4R) and melanocortin-3 (MC3R) receptors with proopiomelanocortin (POMC)-derived peptides (e.g. α-MSH), agouti-related protein (AgRP) and melanocortin-2 receptor accessory protein 2 (MRAP2) is suggested to play critical roles in energy balance of vertebrates. However, evidence on their interaction in birds remains scarce. Our study aims to reveal their interaction in chickens and the results showed that (1) chicken (c-)MC3R and cMC4R expressed in Chinese hamster ovary (CHO) cells can be activated by α-MSH and ACTH1-39 equipotently, monitored by a pGL3-CRE-luciferase reporter system; (2) cMC3R and cMC4R, when co-expressed with cMRAP2 (or cMRAP, a cMRAP2 homolog), show increased sensitivity to ACTH treatment and thus likely act as ACTH-preferring receptors, and the interaction between cMC3R/cMC4R and cMRAP2 was demonstrated by co-immunoprecipitation assay; (3) both cMC3R and cMC4R display constitutive activity when expressed in CHO cells, as monitored by dual-luciferase reporter assay, and cMRAP2 (and cMRAP) can modulate their constitutive activity; (4) AgRP inhibits the constitutive activity of cMC3R/cMC4R, and it also antagonizes ACTH/α-MSH action on cMC4R/cMC3R, indicating that AgRP functions as the inverse agonist and antagonist for both receptors. These findings, together with the co-expression of cMC4R, cMC3R, cMRAP2, cAgRP and cPOMC in chicken hypothalamus detected by quantitative real-time PCR, suggest that within the hypothalamus, α-MSH/ACTH, AgRP and MRAP2 may interact at the MC4R(/MC3R) interface to control energy balance. Furthermore, our data provide novel proof for the involvement of MRAP2 (and MRAP) in fine-tuning the constitutive activity and ligand sensitivity and selectivity of both MC3R and MC4R in vertebrates.