Deciphering the metabolism of LBPA and its function in the endolysosomal system

描述（由申请人提供）：内溶酶体系统缺陷越来越被视为神经退行性疾病的关键病理特征，如阿尔茨海默病（AD）和帕金森病。一种新出现的假设是，这些疾病中发生的慢性内溶酶体缺陷损害了溶酶体的降解能力，导致各种溶酶体货物的异常积累，这些货物通常通过内吞或自噬途径靶向这些细胞器。这些货物不仅包括一系列易于聚集的蛋白质或肽（如β、异常α -突触核蛋白和tau），还包括脂质，如胆固醇和鞘脂。最近，我们的实验室采用了一种称为“脂质组学”的基于系统的方法，使用最先进的质谱法从健康和病变组织中分析了数百种脂质。通过这项技术，我们在Niemann-Pick型C （NPC）小鼠模型中发现了AD患者易感脑区显著的脂质改变，Niemann-Pick型C是一种侵袭性溶酶体储存疾病，与AD有一些共同的致病过程，以及其他载脂蛋白衍生胆固醇异常积累的情况。这种脂质被称为溶双磷脂酸（LBPA）或单酰基甘油磷酸，是一种非典型磷脂，在多泡内体和溶酶体中特别富集，并进一步集中在富含胆固醇的腔内囊泡（ILVs）上。此前已有研究表明，LBPA可促进内溶酶体腔室中ILVs的形成，调节载脂蛋白衍生胆固醇的储存和分布，并通过刺激水解酶促进溶酶体降解。结合这些研究，我们的脂质组学数据不仅确定了LBPA是与内溶酶体功能障碍相关的疾病的候选生物标志物，而且还表明这种磷脂在这些细胞器的生理和病理生理以及载脂蛋白衍生胆固醇的管理中都起着关键作用。不幸的是，目前缺乏工具来了解这种内溶酶体脂质的精确（病理）生理作用，并操纵其水平以评估其治疗潜力。这种障碍背后的主要原因是介导这种难以捉摸的磷脂的合成和降解的酶是未知的。因此，本提案的主要目标是：(i)利用全基因组RNAi筛选和脂质组学相结合，确定LBPA代谢酶，更一般地说，确定正向或负向调节LBPA水平的基因；（ii）评估不同类型的LBPA操作对正常神经元和Npc1突变小鼠神经元中溶酶体功能的影响。我们预计我们的研究将对了解LBPA在溶酶体生理中的作用至关重要，并可能对溶酶体功能障碍相关疾病产生重大影响。

DESCRIPTION (provided by applicant): Defects in the endolysosomal system are increasingly viewed as key pathological features of neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease. An emerging hypothesis is that chronic endolysosomal defects occurring in these disorders compromise the degradative capacity of lysosomes, causing the aberrant accumulation of a variety of lysosomal cargoes that are targeted to these organelles generally through the endocytic or autophagy pathway. These cargoes not only include a range of aggregate-prone proteins or peptides (e.g., Abeta, aberrant alpha-synuclein and tau), but also lipids, such as cholesterol and sphingolipids. Recently, our lab has employed a systems-based approach called "lipidomics" to profile hundreds of lipids from healthy and diseased tissue using state-of-the-art mass spectrometry. With this technology, we identified a striking lipid alteration in vulnerable brain regions from patients with AD, in a mouse model of Niemann-Pick type C (NPC), an aggressive lysosomal storage disorder that shares some pathogenic processes in common with AD, as well as in other instances where apolipoprotein-derived cholesterol is aberrantly accumulating. This lipid is called lysobisphosphatidic acid (LBPA) or bis(monoacylglycero) phosphate, an atypical phospholipid that is specifically enriched in the multivesicular endosomes and lysosomes, where it is further concentrated on cholesterol-rich intralumenal vesicles (ILVs). LBPA has been previously suggested to promote the formation of ILVs in the endolysosomal compartment, to regulate the storage and distribution of apolipoprotein-derived cholesterol, and facilitate lysosomal degradation by stimulating hydrolases. Together with these studies, our lipidomic data not only identify LBPA as a candidate biomarker for disorders associated with endolysosomal dysfunction, but they also suggest a critical role for this phospholipid both in the physiology and the pathophysiology of these organelles as well as in the administration of apolipoprotein-derived cholesterol. Unfortunately, tools are currently lacking to understand the precise (patho)physiological roles of this endolysosomal lipid and manipulate its levels to assess its therapeutic potential. The primary reason behind this roadblock is that the enzymes mediating the synthesis and degradation of this elusive phospholipid are unknown. The main goals of this proposal are thus (i) to identify the LBPA-metabolizing enzymes and more generally, the genes positively or negatively regulating the levels of LBPA using a combination of genome-wide RNAi screen and lipidomics; and (ii) to assess the impact of various types of LBPA manipulations on lysosomal function in normal neurons as well as in neurons derived from Npc1 mutant mice. We anticipate that our studies will be critical to understand the role of LBPA in endolysosomal physiology with potentially major implications for disorders associated with lysosomal dysfunction.