{"title":"依赖于生长因子的 Gαi 磷酸化决定了 G 蛋白偶联受体的典型信号转导。","authors":"Suchismita Roy, Saptarshi Sinha, Ananta James Silas, Majid Ghassemian, Irina Kufareva, Pradipta Ghosh","doi":"10.1126/scisignal.ade8041","DOIUrl":null,"url":null,"abstract":"<div >A long-standing question in the field of signal transduction is how distinct signaling pathways interact with each other to control cell behavior. Growth factor receptors and G protein–coupled receptors (GPCRs) are the two major signaling hubs in eukaryotes. Given that the mechanisms by which they signal independently have been extensively characterized, we investigated how they may cross-talk with each other. Using linear ion trap mass spectrometry and cell-based biophysical, biochemical, and phenotypic assays, we found at least three distinct ways in which epidermal growth factor affected canonical G protein signaling by the G<sub>i</sub>-coupled GPCR CXCR4 through the phosphorylation of Gα<sub>i</sub>. Phosphomimicking mutations in two residues in the α<sub>E</sub> helix of Gα<sub>i</sub> (tyrosine-154/tyrosine-155) suppressed agonist-induced Gα<sub>i</sub> activation while promoting constitutive Gβγ signaling. Phosphomimicking mutations in the P loop (serine-44, serine-47, and threonine-48) suppressed G<sub>i</sub> activation entirely, thus completely segregating growth factor and GPCR pathways. As expected, most of the phosphorylation events appeared to affect intrinsic properties of Gα<sub>i</sub> proteins, including conformational stability, nucleotide binding, and the ability to associate with and to release Gβγ. However, one phosphomimicking mutation, targeting the carboxyl-terminal residue tyrosine-320, promoted mislocalization of Gα<sub>i</sub> from the plasma membrane, a previously uncharacterized mechanism of suppressing GPCR signaling through G protein subcellular compartmentalization. Together, these findings elucidate not only how growth factor and chemokine signals cross-talk through the phosphorylation-dependent modulation of Gα<sub>i</sub> but also how such cross-talk may generate signal diversity.</div>","PeriodicalId":21658,"journal":{"name":"Science Signaling","volume":"17 839","pages":""},"PeriodicalIF":6.7000,"publicationDate":"2024-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Growth factor–dependent phosphorylation of Gαi shapes canonical signaling by G protein–coupled receptors\",\"authors\":\"Suchismita Roy, Saptarshi Sinha, Ananta James Silas, Majid Ghassemian, Irina Kufareva, Pradipta Ghosh\",\"doi\":\"10.1126/scisignal.ade8041\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div >A long-standing question in the field of signal transduction is how distinct signaling pathways interact with each other to control cell behavior. Growth factor receptors and G protein–coupled receptors (GPCRs) are the two major signaling hubs in eukaryotes. Given that the mechanisms by which they signal independently have been extensively characterized, we investigated how they may cross-talk with each other. Using linear ion trap mass spectrometry and cell-based biophysical, biochemical, and phenotypic assays, we found at least three distinct ways in which epidermal growth factor affected canonical G protein signaling by the G<sub>i</sub>-coupled GPCR CXCR4 through the phosphorylation of Gα<sub>i</sub>. Phosphomimicking mutations in two residues in the α<sub>E</sub> helix of Gα<sub>i</sub> (tyrosine-154/tyrosine-155) suppressed agonist-induced Gα<sub>i</sub> activation while promoting constitutive Gβγ signaling. Phosphomimicking mutations in the P loop (serine-44, serine-47, and threonine-48) suppressed G<sub>i</sub> activation entirely, thus completely segregating growth factor and GPCR pathways. As expected, most of the phosphorylation events appeared to affect intrinsic properties of Gα<sub>i</sub> proteins, including conformational stability, nucleotide binding, and the ability to associate with and to release Gβγ. However, one phosphomimicking mutation, targeting the carboxyl-terminal residue tyrosine-320, promoted mislocalization of Gα<sub>i</sub> from the plasma membrane, a previously uncharacterized mechanism of suppressing GPCR signaling through G protein subcellular compartmentalization. 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引用次数: 0
摘要
信号转导领域一个长期存在的问题是,不同的信号通路是如何相互作用来控制细胞行为的。生长因子受体和 G 蛋白偶联受体(GPCR)是真核生物的两大信号枢纽。鉴于它们独立发出信号的机制已被广泛表征,我们研究了它们如何相互交叉。利用线性离子阱质谱法和基于细胞的生物物理、生物化学和表型测定,我们发现表皮生长因子通过 Gαi 磷酸化影响 Gi 偶联 GPCR CXCR4 的典型 G 蛋白信号传导至少有三种不同的方式。Gαi的αE螺旋中两个残基(酪氨酸-154/酪氨酸-155)的磷酸化突变抑制了激动剂诱导的Gαi激活,同时促进了组成型Gβγ信号传导。P 环(丝氨酸-44、丝氨酸-47 和苏氨酸-48)的磷酸化突变完全抑制了 Gi 的激活,从而完全分离了生长因子和 GPCR 途径。不出所料,大多数磷酸化事件似乎都会影响 Gαi 蛋白的固有特性,包括构象稳定性、核苷酸结合以及与 Gβγ 结合和释放 Gβγ 的能力。然而,一个以羧基末端残基酪氨酸-320 为靶点的磷酸模仿突变促进了 Gαi 从质膜上的错误定位,这是一种以前未曾描述过的通过 G 蛋白亚细胞区隔抑制 GPCR 信号转导的机制。总之,这些发现不仅阐明了生长因子和趋化因子信号如何通过磷酸化依赖性调节 Gαi 相互传递,而且还阐明了这种相互传递如何产生信号多样性。
Growth factor–dependent phosphorylation of Gαi shapes canonical signaling by G protein–coupled receptors
A long-standing question in the field of signal transduction is how distinct signaling pathways interact with each other to control cell behavior. Growth factor receptors and G protein–coupled receptors (GPCRs) are the two major signaling hubs in eukaryotes. Given that the mechanisms by which they signal independently have been extensively characterized, we investigated how they may cross-talk with each other. Using linear ion trap mass spectrometry and cell-based biophysical, biochemical, and phenotypic assays, we found at least three distinct ways in which epidermal growth factor affected canonical G protein signaling by the Gi-coupled GPCR CXCR4 through the phosphorylation of Gαi. Phosphomimicking mutations in two residues in the αE helix of Gαi (tyrosine-154/tyrosine-155) suppressed agonist-induced Gαi activation while promoting constitutive Gβγ signaling. Phosphomimicking mutations in the P loop (serine-44, serine-47, and threonine-48) suppressed Gi activation entirely, thus completely segregating growth factor and GPCR pathways. As expected, most of the phosphorylation events appeared to affect intrinsic properties of Gαi proteins, including conformational stability, nucleotide binding, and the ability to associate with and to release Gβγ. However, one phosphomimicking mutation, targeting the carboxyl-terminal residue tyrosine-320, promoted mislocalization of Gαi from the plasma membrane, a previously uncharacterized mechanism of suppressing GPCR signaling through G protein subcellular compartmentalization. Together, these findings elucidate not only how growth factor and chemokine signals cross-talk through the phosphorylation-dependent modulation of Gαi but also how such cross-talk may generate signal diversity.
期刊介绍:
"Science Signaling" is a reputable, peer-reviewed journal dedicated to the exploration of cell communication mechanisms, offering a comprehensive view of the intricate processes that govern cellular regulation. This journal, published weekly online by the American Association for the Advancement of Science (AAAS), is a go-to resource for the latest research in cell signaling and its various facets.
The journal's scope encompasses a broad range of topics, including the study of signaling networks, synthetic biology, systems biology, and the application of these findings in drug discovery. It also delves into the computational and modeling aspects of regulatory pathways, providing insights into how cells communicate and respond to their environment.
In addition to publishing full-length articles that report on groundbreaking research, "Science Signaling" also features reviews that synthesize current knowledge in the field, focus articles that highlight specific areas of interest, and editor-written highlights that draw attention to particularly significant studies. This mix of content ensures that the journal serves as a valuable resource for both researchers and professionals looking to stay abreast of the latest advancements in cell communication science.