Key roles of mono-unsaturated fatty acids in lipid metabolism and lipid-dependent signalling

单不饱和脂肪酸在脂质代谢和脂质依赖性信号传导中的关键作用

• 批准号: RGPIN-2019-06738
• 负责人: Mounier, Catherine
• 金额: $ 3.64万
• 依托单位: Université du Québec à Montréal
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=690006
• 关键词: Key; roles; mono; unsaturated; fatty

Cellular lipid metabolism as well as multiple cell signalling events are impacted by the intracellular levels of monounsaturated fatty acids (MUFA). However, the mechanisms by which MUFA exert these effects remain poorly understood. Our research program aims to elucidate the role of the stearoyl-CoA desaturase 1 (SCD1) enzyme, which produces MUFA, in these cellular events. We use oleate, a major product of the SCD1 enzyme, as a bioactive MUFA in our studies.******OBJECTIVES***1- To determine the role of MUFA in the cellular storage of lipids. We will address the role of SCD1 in intracellular lipid storage, and establish if SCD1 is part of a larger protein complex controlling this process. As oleate preferentially stimulates lipid storage in the form of lipid droplets, we will identify the oleate metabolite responsible for this effect. Further, seipin, a key protein in lipid droplet formation, functionally interacts with SCD1 and, together with SCD1, localizes to the vicinity of lipid droplets; thus, we will examine the role of SCD1 in lipid droplet formation by identifying SCD1 protein interactors under different metabolic states. We will combine information garnered from two cellular models (hepatocytes and yeast) to better understand the contribution of SCD1 to lipid droplet formation.******2- To characterize protein acylation events involving MUFA. We will evaluate the prevalence of MUFA acylation, an emerging type of protein covalent modification, in cellular signalling events. Oleate stimulates lipid synthesis by modulating the activity of SREBP-1, a key lipogenic transcription factor. This modulation of activity likely occurs via direct acylation of SREBP1. Using complementary mass spectrometry analyses, we will identify the lipid moiety on the SREBP-1 protein, as well as its acylated residue(s). We will also use an -alkynyl-biotin pull-down approach to capture palmitoylated and oleylated proteins and identify new MUFA-modified proteins.******3- To determine the impact of MUFA on signalling events associated with cell membranes. We will characterize the impact of oleate on the regulation of signalling events involving lipid second messengers. These events influence cell migration by controlling the formation of cell membrane protrusions. Oleate impacts both cell membrane lipid composition and micro-domain organization, and modulates the activity of membrane signalling complexes. We will characterize the signalling proteins recruited to membrane rafts of oleate-treated cells. Using biosensors, we will also visualize the formation of phospholipase D2 and phosphoinositide 3-kinase products, two enzymes implicated in oleate-stimulated cell migration. We will observe the impact of oleate on the spatial generation of lipid second messengers at the subcellular scale.******Our studies are expected to provide understanding of the molecular mechanisms by which MUFA, particularly oleate, act on signalling pathways regulating cellular homeostasis.**

细胞脂质代谢以及多种细胞信号转导事件受到细胞内单不饱和脂肪酸 (MUFA) 水平的影响。然而，MUFA 发挥这些作用的机制仍然知之甚少。我们的研究计划旨在阐明产生 MUFA 的硬脂酰辅酶 A 去饱和酶 1 (SCD1) 在这些细胞事件中的作用。我们在研究中使用油酸（SCD1 酶的主要产物）作为生物活性 MUFA。 ***** 目标***1- 确定 MUFA 在细胞脂质储存中的作用。我们将探讨 SCD1 在细胞内脂质储存中的作用，并确定 SCD1 是否是控制该过程的更大蛋白质复合物的一部分。由于油酸优先刺激脂滴形式的脂质储存，我们将确定造成这种效应的油酸代谢物。此外，seipin（脂滴形成中的关键蛋白）在功能上与 SCD1 相互作用，并与 SCD1 一起定位于脂滴附近。因此，我们将通过鉴定不同代谢状态下的 SCD1 蛋白相互作用因子来研究 SCD1 在脂滴形成中的作用。我们将结合从两种细胞模型（肝细胞和酵母）获得的信息，以更好地了解 SCD1 对脂滴形成的贡献。********2- 表征涉及 MUFA 的蛋白质酰化事件。我们将评估 MUFA 酰化（一种新兴的蛋白质共价修饰类型）在细胞信号传导事件中的普遍程度。油酸通过调节 SREBP-1（一种关键的脂肪生成转录因子）的活性来刺激脂质合成。这种活性调节可能是通过 SREBP1 的直接酰化发生的。使用互补质谱分析，我们将鉴定 SREBP-1 蛋白上的脂质部分及其酰化残基。我们还将使用炔基生物素下拉方法来捕获棕榈酰化和油化蛋白质，并鉴定新的 MUFA 修饰蛋白质。********3- 确定 MUFA 对与细胞膜相关的信号事件的影响。我们将描述油酸对涉及脂质第二信使的信号事件调节的影响。这些事件通过控制细胞膜突起的形成来影响细胞迁移。油酸影响细胞膜脂质组成和微域组织，并调节膜信号复合物的活性。我们将表征招募到油酸处理细胞膜筏上的信号蛋白。使用生物传感器，我们还将可视化磷脂酶 D2 和磷酸肌醇 3-激酶产物的形成，这两种酶与油酸刺激的细胞迁移有关。我们将观察油酸对亚细胞尺度脂质第二信使空间生成的影响。******我们的研究有望提供对 MUFA（特别是油酸）作用于调节细胞信号通路的分子机制的理解。体内平衡。**