ROLE AND MECHANISMS OF EFFLUX IN MANGANESE HOMEOSTASIS AND DETOXIFICATION

描述(由申请人提供) 锰是一种基本元素，在细胞水平升高时会变得有毒，导致一种无法治愈的神经毒性综合症的发生。到目前为止，关于哺乳动物体内锰的动态平衡和中毒的研究主要集中在锰内流的机制上，但已发现的内流转运体既不是锰的特异性转运体，也不受细胞内锰水平的调节。相反，尽管外流在维持其他金属的动态平衡方面起着重要的作用，但细胞内锰的外流的作用还没有得到充分的研究。研究人员现在已经确定，锰通过摄取进入高尔基体外流，然后分泌，在维持细胞内锰水平和防止在高暴露期间过量积累方面发挥着重要作用。在此基础上，他们的目标是阐明高尔基体锰外流的调控机制，以便更好地了解这一基本过程在锰稳态和锰诱导的神经毒性发展中的作用。研究人员的结果表明，高尔基体摄取锰需要SPCA1，这是一个位于高尔基体内的钙/锰泵，SPCA1在高尔基体和内体之间快速运输。由于离子泵的亚细胞转运调节其活性，他们的首要目标是阐明SPCA1转运在调节锰外流中的作用。调查人员还 研究发现，高尔基体内锰的增加会导致高尔基体蛋白GPP130的快速降解，并且GPP130水平会影响锰的外流和毒性。因此，第二个目的是阐明GPP130调节锰外流和毒性的机制。最后，由于调节锰外流的其他因素仍有待确定，第三个目标是对改变高尔基体对锰的控制的蛋白质进行全基因组RNAi筛选。在屏幕上，改良版本的GPP130将作为高尔基体腔锰的新型传感器。拟议的研究将提供对锰稳态的一个关键但未被探索的方面的机制理解，该方面与锰诱导的神经毒性的病理生物学直接相关。

DESCRIPTION (provided by applicant) Mn is an essential element that becomes toxic at elevated cellular levels leading to the onset of an incurable neurotoxic syndrome. To date, research on Mn homeostasis and toxicosis in mammals has focused on the mechanisms of Mn influx but the identified influx transporters are neither specific for Mn nor regulated by cellular Mn levels. In contrast, the role of efflux of cytosolic Mn is understudied although efflux plays a significant role in maintaining homeostasis of other metals. The investigators have now identified an important role for efflux of Mn via uptake into the Golgi apparatus followed by secretion in maintaining cellular Mn levels and protecting against excess Mn accumulation during elevated exposure. Based on this, their goal is to elucidate the regulatory mechanisms governing Mn efflux by the Golgi to so as to better understand the role of this fundamental process in Mn homeostasis and in the development of Mn-induced neurotoxicity. The investigators' results indicate that uptake of Mn into the Golgi requires SPCA1, a Golgi-localized Ca/Mn pump and that SPCA1 rapidly traffics between the Golgi and endosomes. As subcellular trafficking of ion pumps regulate their activity, their first aim is to elucidate the role of SPCA1 trafficking in regulating Mn efflux. The investigators also discovered that increased intra-Golgi Mn induces rapid degradation of the Golgi protein GPP130 and that GPP130 levels impact Mn efflux and toxicity. Therefore, the second aim is to elucidate the mechanism by which GPP130 regulates Mn efflux and toxicity. Finally, as additional factors regulating Mn efflux remain to be identified, the third aim is to perform a genome wide RNAi screen for proteins that alter control of Mn by the Golgi. In the screen, a modified version of GPP130 will serve as a novel sensor of Golgi lumenal Mn. The proposed studies will provide mechanistic understanding of a crucial but unexplored aspect of Mn homeostasis that is directly relevant to the pathobiology of Mn-induced neurotoxicity.