Meister Jaroslawna, Wanka Lizzy, Perry-Hauser Nicole A, Liu Liu, Iverson Tina M, Gurevich Vsevolod V, Beck-Sickinger Annette G, Kruse Andrew C, Wess Jürgen
Molecular Signaling Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland 20892, United States.
Institute for Clinical Diabetology, German Diabetes Center, Leibniz Institute for Diabetes Research at Heinrich Heine University, Düsseldorf 40225, Germany.
Biochemistry. 2025 Jul 1;64(13):2727-2736. doi: 10.1021/acs.biochem.5c00036. Epub 2025 Jun 9.
The M muscarinic acetylcholine (ACh) receptor (M3R), a prototypic class A biogenic amine G protein-coupled receptor (GPCR), regulates various important functions of the CNS and many of the effects of ACh released from peripheral parasympathetic nerves. Agonist-activated M3Rs preferentially couple to G proteins of the G family but are also able to bind β-arrestins. The recruitment of β-arrestins by the activated M3R disrupts receptor/G protein coupling and promotes M3R internalization, at least in certain cell types. Of note, numerous studies have shown that GPCR-recruited β-arrestins can act as signaling molecules in their own right. These findings raise the question to which extent the recruitment of β-arrestins by activated M3Rs contributes to the physiological responses triggered by M3R activation. A better understanding of the potential physiological relevance of M3R-mediated β-arrestin recruitment may guide the development of so-called biased M3R ligands that preferentially promote either G protein signaling or β-arrestin recruitment. For this reason, the present study aimed to develop two biased mutant M3Rs with opposing functional properties. One of the mutant M3Rs selectively activates G proteins, while the other one preferentially recruits β-arrestins. Here, we report an initial functional characterization of this pair of mutant M3Rs in cultured cells. The future use of these new molecular tools for generating M3R knockin mice or for structural studies may pave the way for the clinical use of biased M3R ligands endowed with high therapeutic efficacy and a favorable side effect profile.
M型毒蕈碱型乙酰胆碱（ACh）受体（M3R）是A类生物胺G蛋白偶联受体（GPCR）的典型代表，可调节中枢神经系统的各种重要功能以及外周副交感神经释放的ACh的许多效应。激动剂激活的M3R优先与G家族的G蛋白偶联，但也能够结合β-抑制蛋白。活化的M3R对β-抑制蛋白的募集会破坏受体/G蛋白偶联，并促进M3R内化，至少在某些细胞类型中是这样。值得注意的是，大量研究表明，GPCR募集的β-抑制蛋白本身可以作为信号分子发挥作用。这些发现提出了一个问题，即活化的M3R对β-抑制蛋白的募集在多大程度上促成了M3R激活引发的生理反应。更好地理解M3R介导的β-抑制蛋白募集的潜在生理相关性，可能会指导所谓的偏向性M3R配体的开发，这些配体优先促进G蛋白信号传导或β-抑制蛋白募集。出于这个原因，本研究旨在开发两种具有相反功能特性的偏向性突变M3R。其中一种突变M3R选择性激活G蛋白，而另一种则优先募集β-抑制蛋白。在此，我们报告了这对突变M3R在培养细胞中的初步功能表征。未来将这些新的分子工具用于生成M3R基因敲入小鼠或进行结构研究，可能为具有高治疗效果和良好副作用谱的偏向性M3R配体的临床应用铺平道路。
