EXPLORING THE ROLE AND THERAPEUTIC POTENTIAL OF RAB GTPASES IN HUNTINGTON'S DISEASE

亨廷顿病(HD)是一种致命的神经退行性疾病，其特征是大脑中脆弱的神经元丢失。这种疾病是由亨廷顿蛋白(HTT)中编码氨基酸谷氨酰胺的重复DNA序列的大小增加引起的。如果HTT蛋白中的谷氨酸数量增加到临界长度以上，它就会错误折叠并聚集在一起形成蛋白质聚集体，扰乱许多重要的细胞过程。值得注意的是，其他基因可以显著改变症状的出现。这表明在人类基因组中有许多潜在的治疗靶点能够显著改变疾病。在患者和疾病模型中的研究表明，突变的HTT(MHTT)影响许多细胞通路，包括细胞内囊泡运输。这是蛋白质和营养物质在细胞不同部分之间移动的重要过程。我们实验室和其他人之前的工作表明，HD中的囊泡运输缺陷至少部分是由于一种名为RAB GTP酶的蛋白质家族所致。这些蛋白质对囊泡运输的几乎每一个方面都是至关重要的，在HD和其他相关疾病中，几个RAB的功能受到损害，包括帕金森氏症。有趣的是，非突变的HTT蛋白可能对一些RAB的正常功能很重要，这可能导致它们在HD中的功能障碍。我们和其他人注意到，在培养的哺乳动物细胞和果蝇HD模型中，三种Rab GTP酶-Rab5、Rab8和Rab11-的表达增加减少了疾病相关症状。虽然这些RAB在HD中的作用已经被探索，但对~60其他哺乳动物RAB的重要性知之甚少。为了解决这个问题，我们对130个哺乳动物RAB及其相关基因进行了系统的筛选，并确定了8个调节哺乳动物细胞中mHTT毒性的基因。在改变mHTT毒性和/或错误折叠的基因的独立筛查中也发现了另外几个RAB。在这项研究中，我们的目标是进一步研究Rab GTP酶在HD中的作用，并探索其治疗潜力。值得注意的是，这将包括通过分析患者来源的样本首次表征RABS在HD患者外周免疫细胞功能障碍中的作用。在HD中，免疫细胞具有高反应性，产生可能导致疾病进展的炎症分子，而RAB有助于这些分子的分泌。了解HD中RAB功能障碍对于确定其疾病和治疗相关性、疾病修改RAB的作用模式和制定治疗方法至关重要。我们将通过调查候选RAB的数量、功能或细胞位置在HD模型和患者样本中是否发生改变，以及这如何影响RAB依赖的过程来解决这一问题。为了验证候选RABS的保护特性，并将其列为进一步研究的优先事项，我们将在果蝇和小鼠HD模型中进行测试，使我们能够研究mHTT和RABS对活的多细胞生物体中细胞之间发生复杂相互作用的影响。有希望的候选者将在与生理相关的人类HD模型细胞中进行测试，包括来自患者的神经元和免疫细胞。改变Rab功能的类药物化合物也将进行测试，并探索靶向它们的替代方法。为了进一步告知潜在的治疗方法并优先考虑候选者，将研究他们改变HD相关表型的机制。我们的初步发现表明，许多候选者增加了mHTT从细胞中的清除，我们将使用其他HD模型和方法来证实这些发现。总之，这项工作将有助于确定Rab GTP酶在HD中的作用，评估它们的治疗潜力，并为治疗策略提供信息。由于RAB功能障碍与几种疾病有关，这些发现也可能具有更广泛的意义。

Huntington's disease (HD) is a fatal neurodegenerative disorder characterised by the loss of vulnerable neurons in the brain. The disease is caused by an increase in the size of a repeated DNA sequence which encodes for the amino acid glutamine in the huntingtin (HTT) protein. If the number of glutamines in the HTT protein increases beyond a critical length, it misfolds and clumps together to form protein aggregates, disrupting many vital cellular processes. Notably, other genes can significantly modify the onset of symptoms. This suggests there are many potential therapeutic targets in the human genome capable of significantly altering disease.Studies in patients and disease models have revealed that mutant HTT (mHTT) affects many cellular pathways, including intracellular vesicle trafficking. This is a vital process for the movement of proteins and nutrients between different parts of the cell. Previous work by our laboratory and others has shown that vesicle trafficking defects in HD are at least partly due to a family of proteins called Rab GTPases. These proteins are vital for nearly every aspect of vesicle trafficking, and the function of several Rabs is impaired in HD and other related disorders, including Parkinson's disease. Interestingly, the non-mutant HTT protein may be important for the normal function of some Rabs, which may contribute to their dysfunction in HD.We and others have noted that increased expression of three Rab GTPases - Rab5, Rab8 and Rab11 - reduces disease-relevant symptoms in cultured mammalian cell and fruit fly models of HD. While the role of these Rabs in HD has been explored, little is known about the importance of the ~60 other mammalian Rabs. To address this question we performed a systematic screen of 130 mammalian Rabs and associated genes and identified 8 that modulated mHTT toxicity in mammalian cells. Several additional Rabs have also been identified in independent screens for genes that alter mHTT toxicity and/or misfolding. In this research proposal we aim to further investigate the role of Rab GTPases in HD and explore their therapeutic potential. Notably, this will include the first characterisation of the role of Rabs in peripheral immune cell dysfunction in HD by analysing patient-derived samples. Immune cells are hyper-reactive in HD, producing inflammatory molecules that may contribute to progression of the disease, and Rabs contribute to the secretion of these molecules.Understanding Rab dysfunction in HD is critical for determining their disease and therapeutic relevance, the mode of action of disease modifying Rabs and for formulating therapeutic approaches. We will address this by investigating whether the amount, function or cellular location of candidate Rabs is altered in HD models and patient samples, and how this impacts upon Rab-dependant processes. To validate the protective properties of candidate Rabs, and prioritise them for further study, they will be tested in fruit fly and mouse HD models, allowing us to study the impact of mHTT and Rabs on the complex interactions that occur between cells in living multicellular organisms. Promising candidates will be tested in physiologically relevant human HD model cells, including neurons and immune cells from patients. Drug-like compounds which alter Rab function will also be tested, and alternative methods for targeting them explored. To further inform potential therapeutic approaches and prioritise candidates the mechanisms by which they modify HD relevant-phenotypes will be studied. Our preliminary findings suggest that many candidates increase the clearance of mHTT from the cell, and we will confirm these findings using additional HD models and approaches. In total, this work will help define the role of Rab GTPases in HD, assess their therapeutic potential and inform therapeutic strategies. As Rab dysfunction has been implicated in several diseases these findings may also have broader significance.