Manganese Metabolism in C. elegans

描述（由申请人提供）：锰（Mn）是一种必需营养素，也是一种有毒的氧化还原金属。因此，必须小心地维持其体内浓度。然而，人们对离子Mn （i-Mn）是如何获得、处理和储存的知之甚少。在一些单细胞生物中，锰已被证明可以弥补抗氧化酶，超氧化物歧化酶的缺乏。在简单的多细胞生物秀丽隐杆线虫（Caenorhabditis elegans）中，我们观察到补充i-Mn增加了对活性氧（ROS）的抗性并延长了寿命。目前尚不清楚i-Mn是如何发挥这种抗氧化活性的。我们的工作假设是，i-Mn主要通过激活叉头相关转录因子DAF-16来发挥这种有益的活性，DAF-16调节秀丽隐杆线虫中几种赋予ROS抗性的基因的表达。此外，我们假设当DAF-16通路被破坏时，i- Mn在体内与合适的配体结合，形成具有ROS中和能力的小分子，可以作为后备防御系统。我们将使用蛋白质组学来确定哪些蛋白质在i-Mn处理下差异表达，并使用daf-16突变体进行表型分析，以测试该转录因子对观察到的i-Mn功能的重要性。我们还将进行功能基因组筛选，以确定i- Mn具有保护性抗氧化功能所需的基因。此外，我们将对蠕虫进行分离，以检查是否在体内形成非蛋白，含锰的低分子量物种，并测试该部分或分离物种是否对秀丽隐杆线虫的ROS提供保护。所提出的研究将为i- Mn的抗氧化活性提供机理细节。这些研究还将确定i-Mn的关键靶点，以及确定参与Mn获取、运输和储存的蛋白质。锰代谢的详细情况可能会出现。此外，拟议的研究将为加州州立大学富勒顿分校的几名本科生和硕士生提供优秀的培训，为他们未来在生物医学科学领域的职业生涯做好更好的准备。

DESCRIPTION (provided by applicant): Manganese (Mn) is both an essential nutrient and a toxic redox metal. Hence its in vivo concentration has to be carefully maintained. Yet, little is known about how ionic Mn (i-Mn) is acquired, handled, and stored. In several unicellular organisms, i-Mn has been shown to compensate for the lack of the antioxidant enzyme, superoxide dismutase. In the simple multi-cellular organism, Caenorhabditis elegans, we have observed that i-Mn supplementation increased resistance to reactive oxygen species (ROS) and extended lifespan. It is unclear how i-Mn exerts this antioxidant-like activity. Our working hypothesis is that i-Mn exerts this beneficial activity primarily through the activation of the Fork head- related transcription factor, DAF-16, which regulates the expression of several genes that confer resistance to ROS in C. elegans. Additionally, we postulate that when DAF-16 pathway is disrupted, i- Mn binds to suitable ligands in vivo to form a small molecule with ROS neutralizing ability, which can act as a backup defense system. We will use proteomics to identify what proteins are differentially expressed with i-Mn treatment and carry out phenotypic analysis using daf-16 mutant to test the importance of this transcription factor for the observed i-Mn function. We will also carry out a functional genomics screen to identify genes required for the protective antioxidant-like function of i- Mn. Additionally, we will fractionate worms to examine if, non-protein, Mn-containing low molecular weight species form in vivo, and test if this fraction or the isolated species provide protection against ROS in C. elegans. The proposed studies will provide mechanistic details for the antioxidant-like activity of i- Mn. These studies will also pinpoint key targets of i-Mn along with identifying proteins involved in Mn acquisition, trafficking and storage. Detailed picture of Mn metabolism is likely to emerge. In addition, the proposed studies will provide outstanding training to several undergraduates and master's level students at California State University, Fullerton, and better prepare them for future careers in Biomedical Sciences. PUBLIC HEALTH RELEVANCE: Free radical mediated damage is associated with over two hundred human diseases and aging. The proposed project utilizes a simple, living, multi-cellular eukaryote, which is comparable to humans, to investigate how a potential free radical scavenger, manganese, works inside a cell. Learning how manganese gets into the cells to exert its beneficial antioxidant-like function is relevant to human health, as it can lead to novel drugs or supplements that could potentially slow the aging process and improve quality of life.