SYNUCLEOPATHIES IN NEURODEGENERATION

描述（由申请人提供）：突触核蛋白家族大量存在于突触前神经元中，并与许多神经退行性疾病状态相关，统称为突触核蛋白病。该家族的一个成员，α-突触核蛋白，作为路易体[LB]的主要成分存在于阿尔茨海默病的LB变体、具有LB的痴呆、散发性帕金森病、多系统萎缩和伴有脑铁积累的神经变性中。α-突触核蛋白的突变体A30 P和A53 T α-突触核蛋白存在于某些遗传形式的PD的LB中。然而，α-突触核蛋白的主要正常功能及其疾病诱导作用的模式都是未知的。了解α-突触核蛋白的分子和功能相关性将有助于了解这种蛋白质的正常和异常活性，这种蛋白质在老年人疾病中发现的氧化应激和炎症增加加速的过程中引起细胞聚集体和LB的形成。 我们提供的初步证据表明，β-突触核蛋白的一个可能的新作用是调节突触前多巴胺转运蛋白[DAT]的活性。DAT活性的这种调节是双峰的，引起转运蛋白活性的增加和正常转运蛋白活性的衰减，如[31 H]多巴胺摄取测定所指示的。DAT功能的调节通过转运蛋白快速运输到质膜和从质膜运输来进行。在DAT活性的这种双重调节中，A30 P和A53 T的作用模式不同于β-突触核蛋白，并且它们也以高度显著的方式彼此不同。 在这个建议中，我们将详细研究的机制，强调这种双峰调节转运活性，使用细胞共转染α-突触核蛋白及其亚型，DAT cDNA，以及在原代培养的神经元。α-突触核蛋白及其A30 P和A53 T突变体的能力，以及参与直接蛋白质：蛋白质复合物形成的结构组分的身份将在正常生长条件下通过一系列研究（包括免疫学、转运蛋白测定、免疫细胞化学和FRET）进行彻底分析。 我们将研究氧化应激在引起α-突触核蛋白的三种变体功能障碍中的作用。 将进行研究以确定α-突触核蛋白/DAT相互作用的变化是否会减少氧化应激和细胞死亡。

DESCRIPTION (provided by applicant): The family of synucleins is abundantly present in presynaptic neurons and is associated with numerous neurodegenerative disease states, collectively termed synucleinopathies. One member of this family, (-synuclein, is present as the major component of Lewy bodies [LB] in LB variant of Alzheimer's disease, dementia with LBs, sporadic Parkinson's disease, multiple system atrophy and neurodegeneration with brain iron accumulation. Mutants of alpha-synuclein, the A30P and A53T alpha-synucleins, are present in LBs of certain genetic forms of PD. However, neither the primary normal function of alpha-synuclein nor its mode of disease inducing action is known. Understanding the molecular and functional correlates of alpha-synuclein would help in the understanding of both the normative, and aberrant activity of this protein that give rise to the formation of fibrillary aggregates and LBs, in a process that is accelerated by increased oxidative stress and inflammation found in diseases of the aged. We provide preliminary evidence to suggest that one possible novel role for (-synuclein is to regulate the activity of the presynaptic dopamine transporter [DAT]. This regulation of DAT activity is bimodal, causing both the increased activity of the transporter and attenuation of normal transporter activity, as is indexed by [31H] dopamine uptake assays. The modulation of DAT function proceeds through rapid trafficking of the transporter to and from the plasma membrane. The mode of action of the A30P and A53T in such dual regulation of DAT activity differs from that of (-synuclein, and they also differ from one another in a highly prominent manner. In this proposal we will study in detail the mechanisms which underline such bimodal regulation of transporter activity, using cells co-transfected with alpha-synuclein and its subtypes, and the DAT cDNA, as well as in primary cultured neurons. The ability of the alpha-synuclein and its A30P and A53T mutants, and the identity of the structural components, which participate in direct protein:protein complex formation will be analyzed thoroughly in normal growth conditional states, through a battery of studies to include, immunology, transporter assays, immunocytochemistry and FRET. We will examine the role of oxidative stress in causing dysfunction of the three variants of alpha-synuclein. Studies will be conducted to determine if changes in alpha-synuclein/DAT interactions will reduce oxidative stress and cell death.