The Role of Desoxysphingoid Bases in HSAN1

描述（由申请人提供）：拟议的跨学科研究针对遗传性感觉和自主神经病变1型（HSAN1）的衰弱性疾病，并基于突变丝氨酸棕榈酰基转移酶（SPT）活性导致两种潜在神经毒性鞘脂的产生的见解。我们首次在突变型HSAN1转基因小鼠和HSAN1患者中发现了这两种非典型去氧鞘样碱基（DSB）的积累。这些研究汇集了生物化学（USUHS制服服务大学）和神经科学和神经遗传学（马萨诸塞州总医院）的专业知识。该项目将(1)研究酵母和哺乳动物细胞中的突变SPT同工酶，(2)对小鼠的非典型鞘脂进行神经毒性研究，(3)评估突变HSAN1转基因小鼠的氨基酸补充情况。这些研究的中心前提是氨基酸底物选择性改变了HSAN1的去氧鞘蛋白碱基和疾病严重程度。密切相关的前提是，人们可以使用行为测试和神经病理检查来评估生化对临床病理表型的影响。我们将首先在酵母和哺乳动物细胞中表达含有已知HSAN1突变的突变SPT异源三聚体（目的1）。每个突变酶对丝氨酸、丙氨酸和甘氨酸的Km和Vmax将使用这些标记的氨基酸来测定。这些实验将使我们能够确定突变是否在浓缩替代底物（丙氨酸和甘氨酸）的能力上存在差异，并为我们在啮齿动物中的研究提供信息。神经毒性研究（目的2）将确定DSB在体内HSAN1周围神经变性中的作用。最后，在突变型HSAN1转基因小鼠中补充丝氨酸与丙氨酸和甘氨酸将使我们能够在体内测试行为和神经病理学（目标3），并为人类临床研究做准备。这些离体和体内研究将使我们对DSB在HSAN1中的合成、降解和神经毒性有更深入的了解。这些实验不仅将阐明对神经代谢的一种新的和基本的见解，而且还将为神经遗传疾病带来一种潜在的新治疗方法。

DESCRIPTION (provided by applicant): The proposed interdisciplinary study addresses the debilitating disorder of hereditary sensory and autonomic neuropathy type 1 (HSAN1) and is based on the insight that mutant serine palmitoyltransferase (SPT) activity leads to production of two potentially neurotoxic sphingolipids. We were the first to identify the accumulation of these two atypical desoxysphingoid bases (DSB) in both mutant HSAN1 transgenic mice and HSAN1 patients. The studies bring together expertise in biochemistry (Uniformed Services University, USUHS) and neuroscience and neurogenetics (Massachusetts General Hospital). The project will (1) investigate mutant SPT isozymes in yeast and mammalian cells, (2) perform neurotoxicity studies of the atypical sphingolipids in mice, and (3) assess amino acid supplementation in mutant HSAN1 transgenic mice. The central premise underlying these investigations is that amino acid substrate selectivity alters desoxysphingoid bases and disease severity in HSAN1. Closely related is the premise that one can use behavioral testing and neuropathological examinations to assess the biochemical impact upon the clinicopathological phenotype. We will begin by expressing mutant SPT heterotrimers containing the known HSAN1 mutations in yeast and in mammalian cells (Aim 1). The Km and Vmax of each mutant enzyme for serine, alanine and glycine will be determined using these labeled amino acids. These experiments will allow us to determine whether the mutations differ in their abilities to condense the alternative substrates (alanine and glycine) and inform our studies in rodents. Neurotoxicity studies (Aim 2) will determine the role of DSB in peripheral neurodegeneration of HSAN1 in vivo. Lastly dietary supplementation of serine versus alanine and glycine in mutant HSAN1 transgenic mice will allow us to test behavior and neuropathology in vivo (Aim 3) and prepare for clinical studies in humans. These ex vivo and in vivo studies will give us insight about the synthesis, degradation and neurotoxicity of DSB in HSAN1. The experiments will not only elucidate a new and fundamental insight into neurometabolism but also lead to a potential new treatment for a neurogenetic disorder.