Non-canonical activation of heterotrimeric G protein signaling in vivo

异源三聚体 G 蛋白信号传导的体内非典型激活

• 批准号: 10461747
• 负责人: Mikel Garcia-Marcos
• 金额: $ 42.61万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-20 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10461747
• 关键词: Affect; Animal Model; Apical; Binding; Biochemical; Biological Assay; Biological Process; Biosensor; Brain; Case Study; Cell physiology; Cells; Congenital Abnormality; Congenital Hydrocephalus; Cues; Cytoplasmic Protein; Data; Development; Disease; Edema; Embryo; Embryonic Development; Encephalopathies; Eukaryota; Family; Family member; G-Protein-Coupled Receptors; G-substrate; GTP-Binding Proteins; Genetic; Goals; Guanine Nucleotide Dissociation Inhibitors; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate; Health; Heterotrimeric GTP-Binding Proteins; Hormones; Human; Hydrocephalus; Hypsarrhythmia; In Vitro; Intercellular Junctions; Knowledge; Measures; Mediating; Mediator of activation protein; Molecular; Mutate; Neurodevelopmental Disorder; Neurotransmitters; Optic Atrophy; Optics; Pharmacology; Physiological Processes; Physiology; Proteins; Regulation; Research; Role; Signal Transduction; Signaling Protein; Specificity; Subgroup; Syndrome; System; Visualization; base; cancer cell; constriction; drug market; genetic regulatory protein; human disease; in vitro activity; in vivo; infancy; innovation; insight; intercellular communication; interest; nervous system development; neurodevelopment; novel; optogenetics; protein activation; prototype; receptor; recruit; synthetic construct; technological innovation; tool

ABSTRACT SIGNIFICANCE: Heterotrimeric G proteins are quintessential mediators of intercellular communication. Defining the molecular mechanisms by which this widespread machinery is regulated is of paramount importance because it impacts a vast range of physiological processes and diseases. This is well exemplified by the ongoing research interest in G protein-coupled receptors (GPCRs), the canonical activators of heterotrimeric G proteins and the targets for >30% of marketed drugs. A much less studied aspect of heterotrimeric G protein regulation, but with far-reaching implications, is their “non-canonical” activation by proteins that are not GPCRs. Among these non-GPCR activators, proteins containing an evolutionarily conserved Gα-Binding-and- Activating (GBA) motif, namely “GBA proteins”, have emerged a key new players in the control of G protein signaling in health and disease. GIV and DAPLE are the prototype members of this family of G protein regulators and both of them are mutated in neurodevelopmental disorders in humans that result in brain birth defects. DAPLE is mutated in non-syndromic congenital hydrocephalus (NSCH) whereas GIV is mutated in Progressive Encephalopathy with edema, Hypsarrhythmia, and Optic atrophy (PEHO)-like syndrome. The cellular and molecular alterations that cause these diseases are poorly understood. BACKGROUND AND GOALS: One of our long-term goals to date has been to identify and characterize a family of cytoplasmic proteins with a GBA motif that activates G protein signaling. Like GPCRs, they have Guanine-nucleotide Exchange Factor (GEF) activity in vitro. Within this context, our goal here is to understand mechanistically how this in vitro GEF activity relates to activation of G proteins and G protein-dependent signaling in living cells, and to establish the importance of this mechanism in vivo. SYNOPSIS OF AIMS: We will (1) dissect how GIV and DAPLE, the two prototype GBAs, affect G protein signaling in cells by using novel chemogenetic platforms and G protein activity biosensors, (2) define how their function is regulated by extrinsic cues by using cell biological assays and novel optogenetic tools, and (3) characterize their impact on neurodevelopment by using animal models. INNOVATION AND IMPACT: Overall, we anticipate that this project will advance the field of heterotrimeric G protein signaling by providing detailed mechanistic insights into a poorly understood aspect of their regulation that impacts human health, and by generating a suite of chemogenetic, optogenetic and biosensor tools to directly manipulate and detect heterotrimeric G protein activity with unprecedented accuracy. The conceptual and technological innovations arising from the successful completion of this project would have a long lasting impact in the field of signal transduction.

抽象的 意义：异三聚体G蛋白是细胞间通信的典型介体。 定义该宽度机械受调节的分子机制至关重要 重要性是因为它会影响广泛的身体过程和疾病。这是很好的例子 通过对G蛋白偶联受体（GPCR）的持续研究兴趣， 杂物中的G蛋白和30％销售药物的靶标。一个较少研究的方面 异三聚合物G蛋白调节，但具有深远的影响，是它们的“非典型”激活 不是GPCR的蛋白质。 在这些非GPCR激活剂中，含有进化的蛋白质构成Gα结合和 - 激活（GBA）主题，即“ GBA蛋白”，已经成为控制G蛋白的主要新参与者 健康和疾病的信号传导。 GIV和DAPLE是该G蛋白家族的原型成员 监管因子和两者都在人类的神经发育障碍中突变，从而导致脑出生 缺陷。在非综合先天性脑积水（NSCH）中，Daple突变为突变，而GIV在 伴有水肿，心律失常和视觉萎缩（PEHO）样综合征的进行性脑病。这 引起这些疾病的细胞和分子改变知之甚少。 背景和目标：迄今为止，我们的长期目标之一就是确定和表征 具有GBA基序的细胞质蛋白家族，可激活G蛋白信号传导。像GPCR一样 鸟嘌呤核苷酸交换因子（GEF）活性在体外。在这种情况下，我们的目标是了解 从机械上讲，这种体外GEF活性与G蛋白的激活和G蛋白依赖性的激活如何 活细胞中的信号传导，并在体内确定该机制的重要性。 目的的概要：我们将（1）剖析两个原型GBA的GIV和Daple如何影响G蛋白 通过使用新型的化学生成平台和G蛋白活性生物传感器在细胞中的信号传导，（2）定义其如何 功能通过使用细胞生物学测定和新型光遗传学工具和（3）来调节外部线索。 通过使用动物模型来表征它们对神经发育的影响。 创新和影响：总的来说，我们预计该项目将推进异三个目标G的领域 蛋白质信号传导通过将详细的机械洞察到其调节的知识方面 这会影响人类健康，并通过产生一套化学遗传，光遗传学和生物传感器工具 直接以前所未有的精度操纵和检测异三聚体G蛋白活性。概念 和由于该项目成功完成而引起的技术创新将有持久的持久 信号转导领域的影响。