Receptor Activity-modifying Protein-directed G Protein Signaling Specificity for the Calcitonin Gene-related Peptide Family of Receptors.
Receptor Activity-modifying Protein-directed G Protein Signaling Specificity for the Calcitonin Gene-related Peptide Family of Receptors.
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DOI：
10.1074/jbc.m116.751362
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发表时间：
2016-10-14
期刊：
影响因子：
--
通讯作者：
Ladds G
中科院分区：
文献类型：
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作者：
Weston C;Winfield I;Harris M;Hodgson R;Shah A;Dowell SJ;Mobarec JC;Woodlock DA;Reynolds CA;Poyner DR;Watkins HA;Ladds G
G蛋白偶联受体（GPCR）的降钙素基因相关肽（CGRP）家族通过降钙素受体样受体（CGRP）与三种受体活性修饰蛋白（RAMP）之一的缔合而形成。众所周知，CGRP、肾上腺髓质素（AM）和中间肽/肾上腺髓质素2（AM 2）这三种肽配体之一的结合可导致Gα s介导的cAMP增加。在这里，我们使用了通过嵌合酵母/Gα亚基将受体活化与细胞生长偶联的修饰酵母菌株和HEK-293细胞来表征不同RAMP和配体组合对该途径的影响。我们不仅证明了与Gαs和Gαq的功能偶联，而且在酵母和HEK-293细胞中鉴定了与Gal信号传导相关的Gαi组分，这在HEK-293 S细胞中不存在。我们发现CGRP受体家族在Gαs、Gαi和Gαq/11通路中显示出配体依赖性和RAMP依赖性信号传导偏好。通过RAMP-GPCR-G蛋白复合物的分子模拟和分子动力学模拟，探讨了RAMP相互作用的背景下，讨论了结果。本研究进一步强调了RAMP对GPCR药理学和一般偏倚的重要性，以及确定选择适当模型系统用于GPCR药理学研究的重要性。
The calcitonin gene-related peptide (CGRP) family of G protein-coupled receptors (GPCRs) is formed through the association of the calcitonin receptor-like receptor (CLR) and one of three receptor activity-modifying proteins (RAMPs). Binding of one of the three peptide ligands, CGRP, adrenomedullin (AM), and intermedin/adrenomedullin 2 (AM2), is well known to result in a Gαs-mediated increase in cAMP. Here we used modified yeast strains that couple receptor activation to cell growth, via chimeric yeast/Gα subunits, and HEK-293 cells to characterize the effect of different RAMP and ligand combinations on this pathway. We not only demonstrate functional couplings to both Gαs and Gαq but also identify a Gαi component to CLR signaling in both yeast and HEK-293 cells, which is absent in HEK-293S cells. We show that the CGRP family of receptors displays both ligand- and RAMP-dependent signaling bias among the Gαs, Gαi, and Gαq/11 pathways. The results are discussed in the context of RAMP interactions probed through molecular modeling and molecular dynamics simulations of the RAMP-GPCR-G protein complexes. This study further highlights the importance of RAMPs to CLR pharmacology and to bias in general, as well as identifying the importance of choosing an appropriate model system for the study of GPCR pharmacology.