How are lipid signalling complexes assembled on membranes

脂质信号复合物如何在膜上组装

• 批准号: RGPIN-2020-04241
• 负责人: Burke, John
• 金额: $ 4.23万
• 依托单位: University of Victoria
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2021
• 资助国家: 加拿大
• 起止时间: 2021-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=739516
• 关键词: How; lipid; signalling; complexes; assembled

Most cellular responses to signals have a common component of regulation arising from the coordinated recruitment of protein complexes to membranes. The coordination of this process is essential to almost every aspect of a cell's life and death. Studying how proteins interact with lipid membranes is extremely challenging by standard biophysical approaches, and has been a major hurdle in understanding signalling at the molecular level. In the last six years of NSERC support (2014-2019), I have made tremendous progress in defining how lipid signalling systems are regulated. The program I am proposing will continue and expand on this strong track record of success to examine the regulation of lipid signalling proteins. The long-term vision of this research program is to understand the molecular details of how large macromolecular assemblies are recruited, assembled, and regulated on lipid membranes. The specific objectives of the research program are as follows: Objectives  1. Define the molecular mechanism of regulation of guanine nucleotide exchange factors (GEFs) on membranes 2. Define how the Phospholipase C (PLC) family of enzymes is regulated on membranes Approach The biological systems under study in this program will be all be approached using a unique synergy of state of the art structural biology approaches, including HDX-MS, Electron Microscopy (EM), and XRC. This approach is unique within Canada, and provides trainees with an exceptional training opportunity. Many of the complexes under study are extremely large (>500 kDa), and are composed of multiple different proteins. The tandem application of these approaches is essential to the study of these complicated systems, and importantly these tools will be applied in a synergistic way, so that the different approaches can inform novel strategies to optimise the likelihood of success in obtaining high resolution structural information. Specifically, this approach allows for the study of high resolution structural information on how these large protein complexes are assembled, together with dynamic studies using HDX-MS to define conformational changes that occur on membrane surfaces. This approach will allow for critical insight into the molecular mechanisms regulating lipid signalling. Impact and significance This research program describes a long term vision of our goals to understand lipid signalling at the molecular level, as well as a set of clear and achievable medium and short term objectives to study lipid signalling systems of extreme complexity. This research will play a vital role in understanding the mechanisms by which lipid signalling complexes are regulated on membranes, which touches on almost all aspects of cell biology. The immediate impact of this proposal on Canada is the training of highly qualified personnel in advanced techniques including molecular biology, HDX-MS, EM, and XRC, which are all in high demand in the biotechnology sector.

大多数细胞对信号的反应都有一个共同的调节成分，由蛋白质复合物协同募集到膜上引起。这个过程的协调对细胞生死的几乎每一个方面都是必不可少的。通过标准的生物物理方法研究蛋白质如何与脂质膜相互作用是极具挑战性的，并且一直是理解分子水平信号传导的主要障碍。在NSERC支持的过去六年中（2014-2019），我在定义脂质信号系统如何调节方面取得了巨大进展。我提议的项目将继续并扩展这一成功的记录，以研究脂质信号蛋白的调节。本研究计划的长期目标是了解大型大分子组装如何在脂质膜上被招募、组装和调节的分子细节。研究计划的具体目标如下：明确鸟嘌呤核苷酸交换因子（GEFs）在膜上调控的分子机制2。定义磷脂酶C （PLC）家族酶是如何在膜上调节的方法在这个项目中研究的生物系统都将使用最先进的结构生物学方法的独特协同作用，包括HDX-MS，电子显微镜（EM）和XRC。这种方法在加拿大是独一无二的，并为学员提供了一个特殊的培训机会。许多正在研究的复合物都非常大（大约500 kDa），并且由多种不同的蛋白质组成。这些方法的串联应用对于这些复杂系统的研究至关重要，重要的是，这些工具将以协同的方式应用，因此不同的方法可以为新的策略提供信息，以优化成功获得高分辨率结构信息的可能性。具体来说，这种方法允许研究这些大蛋白质复合物如何组装的高分辨率结构信息，以及使用HDX-MS定义膜表面发生的构象变化的动态研究。这种方法将允许关键洞察调节脂质信号的分子机制。影响和意义本研究项目描述了我们在分子水平上理解脂质信号传导的长期愿景，以及一套明确和可实现的中期和短期目标，以研究极端复杂的脂质信号传导系统。这项研究将在理解细胞膜上脂质信号复合物的调控机制方面发挥至关重要的作用，这涉及细胞生物学的几乎所有方面。该提案对加拿大的直接影响是培养先进技术的高素质人才，包括分子生物学、HDX-MS、EM和XRC，这些技术在生物技术领域都有很高的需求。