Fine-tuning receptor–G protein activation and signaling

doi:10.1126/science.abc9291

GSTDTAP > 气候变化

DOI	10.1126/science.abc9291
	Fine-tuning receptor–G protein activation and signaling
	Guillaume Lebon
	2020-07-31
发表期刊	Science
出版年	2020
英文摘要	G protein–coupled receptors (GPCRs) are eukaryotic plasma membrane receptors that are organized into four classes in humans: A, B, C, and Frizzled. They internalize extracellular stimuli by activating a common pool of intracellular signaling partners such as the heterotrimeric G proteins (composed of Gα, β, and γ subunits) that subsequently induce an appropriate cellular response. Recent advances in cryo–electron microscopy (cryo-EM) enables challenging structures of GPCR signaling complexes to be solved, providing unprecedented insights about the molecular basis of their signal transduction ([ 1 ][1]). Qiao et al. ([ 2 ][2]) reported two cryo-EM structures of the class B human glucagon receptor (GCGR) Gs and Gi complexes, which helped clarify GCGR G protein selectivity. On page 523 of this issue, Hilger et al. ([ 3 ][3]) report a cryo-EM structure of a GCGR-Gs complex and reveal the effect of conformational changes on GCGR signaling properties. These studies support a common mechanism for class B receptor activation. Activation of G proteins by GPCRs trigger nucleotide exchange and hydrolysis occurring in sequential transition of conformational states from inactive guanosine disphosphate (GDP)-bound forms, to the intermediate nucleotide-free state, then to the active guanosine triphosphate (GTP)–bound G proteins that activate downstream signaling. Accordingly, GPCRs function as guanine nucleotide exchange factors (GEFs). GCGR physiology and signaling are challenging our knowledge of GPCR activation mechanisms. For example, circulating glucagon generated by pancreatic α-cells activates the GCGR and controls glucose homeostasis in the liver ([ 4 ][4]). Once glucagon activates the GCGR-Gs signaling complex in hepatocytes, the Gs protein interacts with adenylate cyclase, which in turn induces production of the second messenger cyclic adenosine monophosphate (cAMP). GCGR has a specific functional signature when considering the amplitude and time scale of glucagon-induced cAMP production compared with other hormones such as epinephrine, which activates the class A GPCR β2-adrenergic receptor (β2AR). It is also common that a GPCR interacts with several G proteins—such as Gs, Gi, and Gq—and GCGR is no exception: It can also activate Gq and Gi proteins ([ 5 ][5]). But how do GPCRs achieve the functional selectivity to activate specific G proteins? The two cryo-EM structures of GCGR-Gs and GCGR-Gi complexes solved by Qiao et al. reveal the large movement of the intracellular tip of transmembrane helix 6 (TM6), which is tilted outward, to accommodate the α5 helix from both Gs and Gi. TM6 movement is a molecular signature of class A and B GPCR active conformations. GCGR TM5 and TM7 undergo a similar motion, but to a lesser extent. This creates a large cavity for the G protein to bind. Conformational changes leading to GCGR activation occur in a different manner than for class A receptors. The conserved residues of the PXXG motif in TM6 are repositioned, and TM6 locally unwinds, which results in a sharp kink that tilts the straight intracellular tip of TM6 away from the receptor core ([ 2 ][2], [ 3 ][3]). Such a kink was reported for several class B GPCRs ([ 6 ][6]), and the superposition of the corresponding receptor signaling complexes clearly illustrates this conserved feature compared with class A receptors, for which there is no kink; instead, TM6 simply bends over, with some degree of variability, depending on the receptor ([ 1 ][1]). The G protein–binding site of type B GPCRs compares well with class A GPCRs coupled to Gs, although it is larger and accommodates a carboxyl-terminal (C-terminal) extremity of the α5 helix from both Gs and Gi (see the figure). There are substantial differences in the sequences of Gs and Gi α5 helices, which is indicative of the requirement of a larger binding site for Gs. The overall contact surface resulting from Gs and Gi subunit engagement with GCGR also differs. This molecular interface is mainly mediated by the α5 helix C-terminal extremity and is larger for Gs, 802 Å2, compared with 551 Å2 for Gi. Another difference is the contribution of the receptor intracellular loop 2 (ICL2) in the GCGR–G protein complexes. The positioning of the Gi αN helix relative to the receptor stresses the ICL2 conformation, making its contribution to the interaction less important for the GCGR-Gi complex, whereas ICL2 makes extended molecular interactions with the αN helix, the β1 strand, and the α5 helix of Gs. The resulting difference in total contact surfaces established between GCGR with Gs and Gi provides an explanation for the lower efficiency of Gi coupling but also for the Gs selectivity over Gi. Disruption of these interfaces by means of site-directed mutagenesis further highlights the importance of the shape and the size of the G protein binding cavity, also stressing the important contribution of ICL2 for G protein selectivity. ![Figure][7] Structural features of G protein activation and selectivity Structural analysis of the class B glucagon receptor (GCGR) reveals the contribution of α5, αN, and transmembrane helix 6 (TM6) helices and intracellular loop 2 (ICL2) conformation to the structural mechanism of G protein selectivity. The activated conformation of GCGR-Gs leads to sustained G protein activation compared with type A β2-adrenergic receptor (β2AR)–Gs signaling. The sharp kink in TM6 of active GCGR illustrates a common mechanism of G protein activation for class B receptors. GRAPHIC: ADAPTED BY A. KITTERMAN/ SCIENCE FROM G. LEBON Once stimulated by glucagon, the GCGR-Gs complex induces a sustained production of cAMP over time compared with β2AR-Gs, suggesting a difference in the kinetics of cAMP accumulation and G protein activation ([ 7 ][8]). Although the structure of the receptor and receptor signaling complexes provide a snapshot of the receptor in action, they do not indicate how the receptor conformation influences G protein activation kinetics. Hilger et al. compare cryo-EM structures of GCGR-Gs to previous β2AR-Gs structures and present an elegant analysis that connects structural differences with differential rates of G protein activation. The cryo-EM structure of GCGR bound to the synthetic glucagon derivative ZP3780 reveals the molecular intricacies involved in the agonist-induced structural changes of TM6, which lead to its intracellular outward shift and the opening of the cavity that allows the G protein to bind. However, ZP3780 is not sufficient to induce the same outward shift of TM6 as observed for β2AR and class A GPCR agonists. Instead, ZP3780 triggers discrete conformational changes in the environment of TM6, stabilizing a GCGR-GDP–bound Gs. This observation correlates with the weak interaction of GS with GCGR ICL2 compared with the β2AR-Gs complex, which likely slows down GDP release and prevents the formation of the fully activated state and nucleotide-free G protein conformation. In addition, in vitro biophysical characterization of purified receptors shows a large difference in GCGR GEF activity, which catalyzes nucleotide exchange 70 times slower than β2AR. Hilger et al. also report slower recruitment of G protein and that GTP binding is three times slower by GCGR, which confirms the reduced rate of G protein activation and nucleotide hydrolysis. It is surprising that the conformation of the TM6 tilt, which is characteristic of the GCGR active conformation, maintains the G protein pocket in an open conformation, even though the G protein has dissociated. This differs from the fast relaxation of the bent TM6 associated with class A GPCRs, which allows them to return to a more energetically stable conformation after G protein activation. Hilger et al. propose that a higher energy barrier separates the GCGR inactive conformation from the intermediate and fully activated states, which potentially accounts for the slower kinetic rate of activation and also for the sustained production of cAMP. In addition to conserved structural features of the activated state, three other class B GPCRs display a slower G protein dissociation rate as compared with that of class A GPCRs, which is indicative of a conserved activation mechanism for class B GPCRs. Our understanding of the complexity and diversity of GPCR activation mechanisms, their coupling selectivity, and all the parameters that may govern their signaling in time and space is only just beginning. GPCR conformational diversity is likely to translate into subtle tuning of intracellular signaling partners such as G proteins, reinforcing the multiplicity of possible biological outputs despite the limited numbers of intracellular partners. In this respect, the composition of the heterotrimeric G protein, namely the β and γ subunits, and other additional regulatory proteins or signaling partners may also affect the G protein activation rate and will require further investigation. More structural information is also needed from different receptor classes to achieve a deeper understanding of their signaling activation mechanism. GPCRs are challenging allosteric machines and are incredibly important drug targets; more than 30% of approved drugs target GPCRs. Class B GPCRs are validated drug targets, and GCGR as well as glucagon–like peptide 1 receptor are important drug targets for type 2 diabetes ([ 8 ][9]). In the future, it will be necessary to combine and integrate structural information with the dynamic properties of receptor conformation and kinetic parameters of GPCR signal transduction when engaging in drug design and the development of new pharmaceuticals. 1. [↵][10]1. J. García-Nafría, 2. C. G. Tate , Mol. Cell. Endocrinol. 488, 1 (2019). [OpenUrl][11][CrossRef][12][PubMed][13] 2. [↵][14]1. A. Qiao et al ., Science 367, 1346 (2020). [OpenUrl][15][Abstract/FREE Full Text][16] 3. [↵][17]1. D. Hilger et al ., Science 369, eaba3373 (2020). [OpenUrl][18][CrossRef][19] 4. [↵][20]1. K. E. Mayo et al ., Pharmacol. Rev. 55, 167 (2003). [OpenUrl][21][Abstract/FREE Full Text][22] 5. [↵][23]1. D. Wootten, 2. L. J. Miller, 3. C. Koole, 4. A. Christopoulos, 5. P. M. Sexton , Chem. Rev. 117, 111 (2017). [OpenUrl][24][CrossRef][25][PubMed][26] 6. [↵][27]1. Y.-L. Liang et al ., Mol. Cell 77, 656 (2020). [OpenUrl][28][CrossRef][29][PubMed][30] 7. [↵][31]1. J. H. Exton, 2. G. A. Robison, 3. E. W. Sutherland, 4. C. R. Park , J. Biol. Chem. 246, 6166 (1971). [OpenUrl][32][Abstract/FREE Full Text][33] 8. [↵][34]1. Y. M. Cho, 2. C. E. Merchant, 3. T. J. Kieffer , Pharmacol. Ther. 135, 247 (2012). [OpenUrl][35][CrossRef][36][PubMed][37] Acknowledgments: This work was supported by the University of Montpellier, Centre National de la Recherche Scientifique, and Institut National de la Santé et de la Recherche Médicale. 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领域	气候变化 ; 资源环境
URL	查看原文
引用统计	被引频次：1[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
条目标识符	http://119.78.100.173/C666/handle/2XK7JSWQ/286856
专题	气候变化资源环境科学
推荐引用方式 GB/T 7714	Guillaume Lebon. Fine-tuning receptor–G protein activation and signaling[J]. Science,2020.
APA	Guillaume Lebon.(2020).Fine-tuning receptor–G protein activation and signaling.Science.
MLA	Guillaume Lebon."Fine-tuning receptor–G protein activation and signaling".Science (2020).