Abstract
G protein-coupled receptors (GPCRs) are regulated by various downstream proteins, of which the melanocortin receptor accessory protein 1 (MRAP1) is closely involved in the regulation of melanocortin receptor 2 (MC2R). Assisted by MRAP1, MC2R responds to adrenocorticotropic hormone (ACTH) and stimulates glucocorticoid biogenesis and cortisol secretion. MC2R activation plays an essential role in the hypothalamic-pituitary-adrenal (HPA) axis that regulates stress response, while its dysfunction causes glucocorticoid insufficiency- or cortisol excess-associated disorders. Here, we present a cryo-electron microscopy (cryo-EM) structure of the ACTH-bound MC2R–Gs–MRAP1 complex. Our structure, together with mutagenesis analysis, reveals a unique sharp kink at the extracellular region of MRAP1 and the ‘seat-belt’ effect of MRAP1 on stabilizing ACTH binding and MC2R activation. Mechanisms of ACTH recognition by MC2R and receptor activation are also demonstrated. These findings deepen our understanding of GPCR regulation by accessory proteins and provide valuable insights into the ab initio design of therapeutic agents targeting MC2R.
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Data availability
The corresponding coordinates and cryo-EM density map have been deposited in the Protein Data Bank (http://www.rcsb.org/pdb) with code 8GY7, and in EMDB (http://www.ebi.ac.uk/pdbe/emdb/) with code EMD-34371, respectively.
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Acknowledgements
The cryo-EM data of the ACTH–MC2R–Gs–MRAP1 complex were collected at the Advanced Center for Electron Microscopy, Shanghai Institute of Materia Medica (SIMM). We thank all the staff at the cryo-EM facilities for their technical support. This work was partially supported by the National Natural Science Foundation (32171187 to Y.J., 82121005 to Y.J. and H.E.X., 32130022 to H.E.X., 81872915 to M.-W.W., 81773792 and 81973373 to D.Y.); the Ministry of Science and Technology (China) grants (2018YFA0507002 to H.E.X. and M.-W.W.); the Shanghai Municipal Science and Technology Major Project (2019SHZDZX02 to H.E.X.); Shanghai Municipal Science and Technology Major Project (H.E.X.); the CAS Strategic Priority Research Program (XDB37030103 to H.E.X.).
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P.L. designed the expression constructs, purified the MC2R complex protein, prepared the final samples for negative stain and cryo-EM data collection, conducted functional studies, and prepared figures and manuscript draft; W.F. conducted functional studies with the help of A.D., X. Chen, and X. Cai, supervised by M.-W.W. and D.Y.; S.M. participated in the preparation of constructs and expression screening; K.W. and Q.Y. participated in cryo-EM data acquisition; M.-W.W. participated in manuscript preparation; Y.J. and H.E.X. conceived and supervised the project, analyzed the structures, and wrote the manuscript with inputs from all authors.
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Luo, P., Feng, W., Ma, S. et al. Structural basis of signaling regulation of the human melanocortin-2 receptor by MRAP1. Cell Res 33, 46–54 (2023). https://doi.org/10.1038/s41422-022-00751-6
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DOI: https://doi.org/10.1038/s41422-022-00751-6
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