Understanding and Manipulating G Protein α Subunit and Phospholipase C Signaling Networks

了解和操作 G 蛋白 α 亚基和磷脂酶 C 信号网络

• 批准号: 10621415
• 负责人: Alan V. Smrcka
• 金额: $ 53.64万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10621415
• 关键词: Adenylate Cyclase; Adrenergic Agents; Adrenergic Receptor; Area; Cardiac Myocytes; Cells; Development; Functional disorder; Funding; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Protein alpha Subunits; GTP-Binding Proteins; Gene Expression; Genes; Golgi Apparatus; Heart Hypertrophy; Heart failure; Hydrolysis; Individual; Label; Laboratories; Ligands; Mass Spectrum Analysis; Mediating; Membrane; Methods; Molecular; Pathway interactions; Pharmaceutical Preparations; Phosphatidylinositols; Phospholipase C; Physiology; Play; Process; Proteomics; Receptor Signaling; Recycling; Regulation; Rest; Role; Second Messenger Systems; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Testing; Tissues; beta-adrenergic receptor; career; chromatin remodeling; experimental study; follow-up; heart function; in vivo; interest; novel; novel therapeutics; protein protein interaction; receptor; response; success; targeted treatment

My laboratory is interested signal transduction by G protein-coupled receptors (GPCRs) with a specific focus on the second messenger pathways downstream and how they drive physiology and pathophysiology in specific cells and tissues. GPCRs signaling pathways are ubiquitous and conserved, with cellular responses determined by 1) the specific GPCRs and G proteins expressed in the cell and 2) how the cells decode signals generated by these receptors. My career has been focused on uncovering novel cellular signal transduction mechanisms at a cellular and molecular level that move beyond a canonical GPCR paradigm where individual GPCRs simply couple to 3 major signal transduction pathways, phospholipase C activation, adenylyl cyclase regulation, and RhoGEF stimulation, and the cells do the rest. In this renewal application we propose to capitalize on advances made in the previous funding in two general areas 1) Proteomic analysis of G protein α subunit-mediated signal transduction pathways and; 2) intracellular signaling by β-adrenergic receptors (βARs) and phospholipase C in cardiac myocytes. The first project is based on the striking success of our proteomic screens using proximity labeling mass spectrometry that maintains cell context while identifying protein-protein interactions. We propose to characterize two novel G protein targets from these screens that play roles in regulation of chromatin remodeling and gene expression through mechanisms that would be unprecedented downstream of GPCRs. In addition, we propose to leverage this method to identification of new signal pathways mediated by G proteins as GPCRs move through the endocytic and recycling pathways. Intracellular signaling by GPCRs is a new and emerging paradigm in the field but much less is known about intracellular G protein and effector engagement by internal GPCRs. The second direction will be to follow up our finding that phospholipase C activity at the Golgi apparatus is regulated by intracellular Golgi localized β1-adrenergic receptors (β1ARs). The proposed experiments will test the hypothesis that intracellular β1ARs regulate a unique subset of genes in cardiac myocytes related to cardiac hypertrophy through regulation of phosphoinositide hydrolysis at the Golgi apparatus. We will determine what signaling pathways are regulated by Golgi β1ARs beyond the Epac-PLCε- PKD signaling pathway that we have previously described. Finally, we will investigate the mechanisms for pathways downstream of β2ARs that oppose hypertrophic β1AR signaling at the Golgi. This is exciting because β1AR signaling in vivo is pro-hypertrophic while β2AR signaling is protective and will uncover a novel mechanism for protective β2AR signaling. Overall, these experiments will reveal novel signaling mechanisms that have implications for therapies that target GPCRs.

我的实验室对 G 蛋白偶联受体 (GPCR) 的信号转导感兴趣，特别关注 第二信使通路下游以及它们如何驱动特定的生理学和病理生理学 GPCR 信号通路普遍存在且保守，细胞反应是确定的。 1) 细胞中表达的特定 GPCR 和 G 蛋白，以及 2) 细胞如何解码生成的信号 我的职业生涯一直致力于发现新的细胞信号转导机制。 在细胞和分子水平上，超越了规范的 GPCR 范式，在规范的 GPCR 范式中，单个 GPCR 简单地 耦合到 3 个主要信号转导途径：磷脂酶 C 激活、腺苷酸环化酶调节和 RhoGEF 刺激，细胞完成剩下的工作。在这个更新应用中，我们建议利用先进技术。 之前的资助主要在两个领域进行：1）G蛋白α亚基介导的信号的蛋白质组学分析 转导途径；2) β-肾上腺素受体 (βAR) 和磷脂酶 C 的细胞内信号传导 第一个项目基于我们使用邻近法进行蛋白质组学筛选的巨大成功。 我们建议使用标记质谱法来维持细胞背景，同时识别蛋白质-蛋白质相互作用。 从这些筛选中表征两个在染色质调节中发挥作用的新型 G 蛋白靶点 通过 GPCR 下游前所未有的机制进行重塑和基因表达。 此外，我们建议利用这种方法来鉴定由 G 蛋白介导的新信号通路，如 GPCR 通过内吞和再循环途径进行 GPCR 的细胞内信号传导是一种新的途径。 该领域正在出现新兴范例，但人们对细胞内 G 蛋白和效应子参与知之甚少。 第二个方向是跟进我们对高尔基体磷脂酶 C 活性的发现。 该装置由细胞内高尔基体局部 β1 肾上腺素受体 (β1AR) 调节。 实验将检验细胞内 β1AR 调节心脏中独特的基因子集的假设 通过调节高尔基体磷酸肌醇水解与心肌肥厚相关的心肌细胞 我们将确定 Epac-PLCε- 之外的高尔基体 β1AR 调节哪些信号通路。 我们之前描述过的 PKD 信号通路最后，我们将研究其机制。 β2AR 的下游通路对抗高尔基体肥大的 β1AR 信号传导，这是令人兴奋的。 体内 β1AR 信号传导具有促肥大作用，而 β2AR 信号传导具有保护作用，将揭示一种新机制 总的来说，这些实验将揭示具有保护作用的β2AR信号传导机制。 对针对 GPCR 的疗法的影响。

项目成果

Stabilization of interdomain interactions in G protein α subunits as a determinant of Gαi subtype signaling specificity.

G 蛋白 α 亚基中域间相互作用的稳定作为 Gαi 亚型信号传导特异性的决定因素。

• DOI: 10.1016/j.jbc.2024.107211
• 发表时间: 2024
• 期刊: The Journal of biological chemistry
• 作者: Lefevre,TylerJ;Wei,Wenyuan;Mukhaleva,Elizaveta;MedaVenkata,SaiPranathi;Chandan,NaincyR;Abraham,Saji;Li,Yong;Dessauer,CarmenW;Vaidehi,Nagarajan;Smrcka,AlanV
• 通讯作者: Smrcka,AlanV

Subcellular β-Adrenergic Receptor Signaling in Cardiac Physiology and Disease.

• DOI: 10.1097/fjc.0000000000001324
• 发表时间: 2022-09-01
• 期刊: JOURNAL OF CARDIOVASCULAR PHARMACOLOGY
• 影响因子: 3
• 作者: Wei, Wenhui;Smrcka, Alan, V
• 通讯作者: Smrcka, Alan, V

β-arrestin mediates communication between plasma membrane and intracellular GPCRs to regulate signaling.

• DOI: 10.1038/s42003-020-01510-2
• 发表时间: 2020-12-18
• 期刊: Communications biology
• 影响因子: 5.9
• 作者: DeNies MS;Smrcka AV;Schnell S;Liu AP
• 通讯作者: Liu AP