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（GPCRs）的信号转导感兴趣，特别关注 第二信使通路下游，以及它们如何驱动生理学和病理生理学， 细胞和组织。GPCR信号通路是普遍存在的和保守的，细胞反应确定 通过1）细胞中表达的特异性GPCR和G蛋白以及2）细胞如何解码产生的信号 这些受体。我的职业生涯一直专注于发现新的细胞信号转导机制 在细胞和分子水平上，超越了典型的GPCR范式，其中单个GPCR仅仅是 偶联3个主要信号转导途径，磷脂酶C激活，腺苷酸环化酶调节， RhoGEF刺激，细胞做剩下的。在这份续约申请中，我们建议利用预付款 在以往的资助中主要做了两个方面的工作：1）G蛋白α亚基介导信号的蛋白质组学分析 转导途径; 2）β-肾上腺素能受体（β AR）和磷脂酶C的细胞内信号转导， 心肌细胞第一个项目是基于我们的蛋白质组学筛选的惊人成功， 标记质谱，保持细胞环境，同时识别蛋白质-蛋白质相互作用。我们提出 从这些筛选中表征两个在染色质调节中起作用的新G蛋白靶点， 重塑和基因表达的机制，这将是前所未有的下游GPCR。在 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， GPCR通过内吞和再循环途径移动。通过GPCR的细胞内信号传导是一种新的， 但对细胞内G蛋白和效应子的参与知之甚少， 内部GPCR。第二个方向将是跟进我们的发现，磷脂酶C的活性在高尔基体 细胞器受细胞内高尔基体定位的β1肾上腺素能受体（β1ARs）调节。拟议 实验将检验细胞内β 1AR调节心脏中一个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大相关的心肌细胞 设备.我们将确定高尔基体β 1 AR在Epac-PLCε-受体之外调节哪些信号通路。 PKD信号通路，我们以前已经描述过。最后，我们将研究 β 2 AR下游通路，其对抗高尔基体处的肥大β 1 AR信号传导。这很令人兴奋，因为 体内β 1 AR信号是促肥大的，而β 2 AR信号是保护性的，这将揭示一种新的机制 保护性β2AR信号。总的来说，这些实验将揭示新的信号机制， 对靶向GPCR的治疗的影响。

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