Key Laboratory of Bio-resources and Eco-environment of Ministry of Education, College of Life Sciences, Sichuan University, Chengdu 610065, China.
Key Laboratory of Bio-resources and Eco-environment of Ministry of Education, College of Life Sciences, Sichuan University, Chengdu 610065, China.
Domest Anim Endocrinol. 2020 Jul;72:106435. doi: 10.1016/j.domaniend.2020.106435. Epub 2020 Jan 11.
The 2 structurally related peptides, neuropeptide FF (NPFF) and neuropeptide AF (NPAF), are encoded by the NPFF gene and have been identified as neuromodulators that regulate nociception and opiate-mediated analgesia via NPFF receptor (NPFFR2) in mammals. However, little is known about these 2 peptides in birds. In this study, we examined the structure, tissue expression profile, and functionality of NPAF and NPFF in chickens. Our results showed that: 1) unlike mammalian NPFF, NPFF from chicken and other avian species is predicted to produce a single bioactive NPAF peptide, whereas the putative avian NPFF peptide likely lacks activity due to the absence of functional RFamide motif at its C-terminus; 2) synthetic chicken (c-) NPAF can potently activate cNPFFR2 (and not cNPFFR1) expressed in HEK293 cells, as monitored by 3 cell-based luciferase reporter systems, indicating that cNPAF is a potent ligand for cNPFFR2, which activation could decrease intracellular cAMP levels and stimulate the MAPK/ERK signaling cascade; interestingly, gonadotropin-inhibitory hormone, a peptide sharing high structural similarity to NPAF, could specifically activate cNPFFR1 (but not cNPFFR2); 3) Quantitative real-time PCR revealed that cNPFF mRNA is widely expressed in chicken tissues with the highest level detected in the hypothalamus, whereas cNPFFR2 is expressed in all tissues examined with the highest level noted in the hypothalamus and anterior pituitary. Taken together, our data reveal that avian NPFF encodes a single bioactive NPAF peptide, which preferentially activates NPFFR2, and provides insights into potential structural and functional changes of NPFF-derived peptides during vertebrate evolution.
这 2 种结构相关的肽，神经肽 FF（NPFF）和神经肽 AF（NPAF），由 NPFF 基因编码，已被鉴定为调节哺乳动物痛觉和阿片类药物介导的镇痛的神经调节剂，通过 NPFF 受体（NPFFR2）发挥作用。然而，关于鸟类的这 2 种肽知之甚少。在这项研究中，我们研究了 NPAF 和 NPFF 在鸡中的结构、组织表达谱和功能。我们的结果表明：1）与哺乳动物 NPFF 不同，来自鸡和其他鸟类的 NPFF 预计会产生一种单一的生物活性 NPAF 肽，而假定的鸟类 NPFF 肽可能由于其 C 末端缺乏功能性 RFamide 基序而缺乏活性；2）合成的鸡（c-）NPAF 可以强烈激活在 HEK293 细胞中表达的 cNPFFR2（而不是 cNPFFR1），如 3 种基于细胞的荧光素酶报告系统所监测到的，表明 cNPAF 是 cNPFFR2 的有效配体，其激活可以降低细胞内 cAMP 水平并刺激 MAPK/ERK 信号级联；有趣的是，促性腺激素抑制激素，一种与 NPAF 具有高度结构相似性的肽，可以特异性地激活 cNPFFR1（而不是 cNPFFR2）；3）实时定量 PCR 显示，cNPFF mRNA 在鸡组织中广泛表达，在下丘脑检测到的水平最高，而 cNPFFR2 在所有检测到的组织中表达，在下丘脑和垂体前叶的水平最高。总之，我们的数据揭示了禽类 NPFF 编码一种单一的生物活性 NPAF 肽，其优先激活 NPFFR2，并为脊椎动物进化过程中 NPFF 衍生肽的潜在结构和功能变化提供了见解。
