alpha-synuclein and protein degradation

描述（由申请人提供）：帕金森病（PD）是美国和其他地区老龄化人口的主要健康负担。在罕见家族中发现导致该病的基因缺陷可能会对更常见的散发形式的病理生理学产生新的见解。与帕金森病相关的一个特殊基因是α -突触核蛋白（ASYN），因为它的蛋白产物是路易体的主要成分。此外，ASYN基因座的增殖足以导致人类PD。因此，阐明ASYN的正常和异常功能及其调控方式，可能有助于更好地了解PD的病理生理学和开发新的治疗方法。我们之前已经发现突变型ASYN可以通过溶酶体和泛素-蛋白酶体系统破坏蛋白质降解。蛋白质处理缺陷也可能发生在其他遗传或散发形式的PD中，并可能形成该疾病的共同病理生理基础。我们提出，对溶酶体的影响可能是由突变体ASYN对伴侣介导的自噬（CMA）途径的抑制介导的，CMA通常会降解ASYN。然而，这些结果主要是在人工环境中取得的，并且ASYN的降解及其对蛋白水解系统的影响是有争议的。我们建议首先通过操纵CMA受体Lamp2a及其限速步骤，并通过检测ASYN不靶向CMA途径时的降解，在神经细胞中确定ASYN被CMA降解。不影响CMA的突变型ASYN也将被检查其相对毒性作用，以确定ASYN毒性是否部分由CMA阻断介导。我们最近发现ASYN可以分泌并发挥毒性作用，我们将研究这种可能与生理相关的毒性是否与神经元细胞中UPS和/或溶酶体功能的抑制有关。最后，我们将研究ASYN对神经细胞系中蛋白质降解系统的影响是否适用于原代神经细胞培养。这些研究将有助于破译ASYN水平的翻译后调控，这对于其稳态水平及其对神经元生理的影响至关重要，并且它对蛋白质降解途径的影响可能是其致病作用的关键。

DESCRIPTION (provided by applicant): Parkinson's Disease (PD) is a major health burden on the ageing population in the United States and elsewhere. The identification of gene defects that cause the disease in rare families may lead to new insights into the pathophysiology of the more common sporadic forms. One particular gene linked to PD that is especially relevant to the sporadic disease is alpha-synuclein (ASYN), as its protein product is the major component of Lewy Bodies. Furthermore, multiplication of the ASYN locus is sufficient to cause PD in humans. Therefore, elucidating the normal and aberrant functions of ASYN, as well as the manner of its regulation, may lead to a better understanding of the pathophsyiology of PD and the development of novel therapies. We have previously found that mutant ASYN can impair protein degradation through the lysosomes and the ubiquitin-proteasome system. Defects of protein handling may also occur in other genetic or sporadic forms of PD, and may form a common pathophysiological basis for this disorder. We have proposed that the impact on the lysosomes may be mediated by an inhibition by mutant ASYN of the pathway of Chaperone-Mediated Autophagy (CMA), which normally degrades ASYN. However, these results were largely achieved in an artificial setting, and the degradation of ASYN and its impact on proteolytic systems is controversial. We propose to first establish that ASYN is degraded by CMA in neuronal cells, by manipulating Lamp2a, the receptor for CMA and its rate-limiting step, and by examining the degradation of ASYN when it is not targeted to the CMA pathway. Mutant ASYNs that do not affect CMA will also be examined for their relative toxic effects, to ascertain whether ASYN toxicity is mediated in part by CMA blockade. We have recently found that ASYN can be secreted and exert toxic effects, and we will examine whether such toxicity, which may be physiologically relevant, is related to the inhibition of UPS and/or lysosomal function in neuronal cells. Lastly, we will examine whether the effects of ASYN on protein degradation systems in neuronal cell lines are applicable in primary neuronal cell cultures. These studies will help to decipher the post-translational regulation of ASYN levels, which is critical for its steady-state levels and its impact on neuronal physiology, and its effects on protein degradation pathways, which may be critical for its pathogenic effects.