Lipotoxic Protective Response of the Endoplasmic Reticulum

内质网的脂毒性保护反应

• 批准号: 10706013
• 负责人: ROBERT V FARESE
• 金额: $ 19.19万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10706013
• 关键词: Lipotoxic; Protective; Response; Endoplasmic; Reticulum

Lipid homeostasis is crucially important for cells, and disruptions in this balance can lead to diseases of lipid overload, such as obesity-related disorders. Lipid homeostasis is mainly coordinated in the endoplasmic reticulum (ER), the largest membrane system in cells. The ER is a well-recognized location for synthesis of membrane and secreted proteins, processes that are safeguarded by the “unfolded protein response” (UPR). Recent studies suggest that a complementary and interrelated pathway, which we term the “lipotoxic protective response” (LPR), acts in parallel to maintain ER lipid homeostasis. In contrast to the UPR, however, knowledge of the molecular aspects of the LPR remain rudimentary. Here we describe studies to overcome this knowledge gap through investigations through our recent identification of the mammalian FIT2 protein as a key guardian of ER lipid homeostasis. FIT2 has been a mysterious ER protein, of crucial importance for cell health and function, that was implicated in ER homeostasis and lipid metabolism but lacked an identified function. After years of effort and a biochemical tour de force, we have discovered that FIT2 is an acyl-CoA diphosphatase enzyme that catabolizes fatty acyl-CoAs, the activated forms of fatty acids, in the ER. Our preliminary data indicate that this activity may be localized on the luminal leaflet and is crucially important for cell health and viability. Absence of FIT2 triggers ER stress and, in a mouse model we have generated, liver injury. Our findings break new ground and raise many important questions about FIT2 and the LPR. Here we propose to answer these questions by using an interdisciplinary approach. Aim 1 will determine mechanisms of the LPR at the molecular level by deciphering the biochemical mechanism for FIT2’s catabolism of fatty acyl-CoA in the ER. We will combine biochemical and structural biology approaches to answer: What is the enzyme’s catalytic mechanism? Is FIT2 active towards ER luminal substrates? Is its activity regulated by lipid metabolites (e.g., diacylglycerol, DAG)? What is the fate of the metabolites generated by FIT2 activity? Aim 2 will determine mechanisms of the LPR at the cellular level by elucidating how FIT2 and the LPR are integrated with other ER stress protection pathways. Specifically, we will address: How does FIT2 mechanistically trigger the UPR? Does the mammalian UPR pathway depend on FIT2 activity? Can the interdependency of the FIT2/LPR and the UPR be exploited for therapeutic purposes in cancer, for example those with high demands on ER function, such hepatocellular carcinoma and multiple myeloma? Aim 3 will determine mechanisms of the LPR at the physiological level by determining how FIT2 maintains lipid homeostasis in mammalian liver. We have generated mice lacking FIT2 in hepatocytes, and preliminary studies indicate these mice have ER stress, increases in hepatocyte lipid storage, defects in lipid secretion, and signs of liver disease. We will mechanistically dissect this phenotype to elucidate how FIT2 is required in cellular lipid metabolism. Completion of these aims will reveal how a fast, enzymatic mechanism functions in the LPR to protect cells and maintain lipid homeostasis. It will also shed light on the pathogenesis of human disease, such as the rare human disorder of FIT2 deficiency and more common metabolic diseases involving ER lipid metabolism, with implications for both liver steatosis and cancers that rely heavily of the ER (such as liver cancer and multiple myeloma).

脂质稳态对细胞至关重要，这种平衡的破坏可导致脂质过载疾病， 例如肥胖相关的疾病。脂质稳态主要在内质网（ER）中协调，内质网是最大的内质网。 细胞膜系统。ER是公认的合成膜和分泌蛋白的位置， 这一过程受到“未折叠蛋白质反应”（UPR）的保护。最近的研究表明， 和相关的途径，我们称之为“脂毒性保护反应”（LPR），平行作用，以维持ER脂质 体内平衡然而，与普遍定期审议相比，对LPR的分子方面的了解仍然是初步的。这里 我们描述的研究，以克服这一知识差距，通过调查，通过我们最近的鉴定， 哺乳动物FIT 2蛋白作为ER脂质稳态的关键监护人。FIT 2是一种神秘的ER蛋白， 对细胞健康和功能的重要性，它与ER稳态和脂质代谢有关，但缺乏 识别功能。经过多年的努力和生化绝技，我们发现FIT 2是一种酰基辅酶A， 在ER中分解代谢脂肪酰辅酶A（脂肪酸的活化形式）的二磷酸酶。我们的初步数据 表明这种活性可能位于管腔小叶上，并且对细胞健康和活力至关重要。 缺乏FIT 2会触发内质网应激，在我们生成的小鼠模型中，会引发肝损伤。我们的发现开辟了新天地 并提出了许多关于FIT 2和LPR的重要问题。在这里，我们建议通过使用一个 跨学科方法。目的1将通过破译分子水平的LPR来确定LPR的机制， FIT 2在ER中催化脂肪酰辅酶A的生化机制。我们将结合联合收割机的生化和结构 生物学的方法来回答：酶的催化机制是什么？FIT 2对ER管腔底物有活性吗？ 其活性是否受脂质代谢物（例如，甘油二酯（diacylglycerol，DAG）？由细菌产生的代谢物的命运是什么 FIT 2活动？目的2将通过阐明FIT 2和LPR如何在细胞水平上确定LPR的机制。 与其他ER应激保护途径相结合。具体而言，我们将解决：FIT 2如何机械地触发 普遍定期审议？哺乳动物的UPR通路依赖于FIT 2活性吗？FIT 2/LPR和 UPR可用于癌症的治疗目的，例如对ER功能有高要求的癌症，如 肝细胞癌和多发性骨髓瘤目的3将从生理水平上确定LPR的机制 通过确定FIT 2如何维持哺乳动物肝脏中的脂质稳态。我们已经产生了缺乏FIT 2的小鼠， 初步研究表明，这些小鼠具有ER应激、肝细胞脂质储存增加、 脂质分泌和肝脏疾病的迹象。我们将机械地剖析这种表型，以阐明FIT 2是如何被激活的。 是细胞脂质代谢所必需的。这些目标的完成将揭示一种快速的酶机制是如何发挥作用的， LPR保护细胞和维持脂质稳态。它还将揭示人类疾病的发病机制， 作为罕见的人类FIT 2缺乏症和更常见的涉及ER脂质代谢的代谢疾病， 这对肝脏脂肪变性和严重依赖ER的癌症（如肝癌和多发性骨髓瘤）都有意义。