Manganese Metabolism in C. elegans

说明(申请人提供)：锰既是一种必需的营养物质，也是一种有毒的氧化还原金属。因此，必须小心地保持其体内浓度。然而，关于离子锰(I-Mn)是如何获得、处理和储存的，人们知之甚少。在一些单细胞生物体中，I-Mn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体秀丽线虫中，我们观察到I-Mn的补充增加了对活性氧(ROS)的抵抗，延长了寿命。目前尚不清楚I-Mn是如何发挥这种类似抗氧化剂的活性的。我们的工作假设是，I-Mn主要通过激活与叉头相关的转录因子DAF-16来发挥这种有益的活性，DAF-16调节线虫中几个赋予ROS抗性的基因的表达。此外，我们推测当DAF-16途径被破坏时，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，作为后备防御系统。我们将使用蛋白质组学来确定I-mN处理后哪些蛋白质有差异表达，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学筛选，以确定I-mN保护性抗氧化剂样功能所需的基因。此外，我们将对蠕虫进行分离，以检查体内是否形成非蛋白质、含锰的低分子量物种，并测试该部分或分离的物种是否对线虫的ROS具有保护作用。拟议的研究将为I-MN的抗氧化剂样活性提供机制细节。这些研究还将确定I-MN的关键靶点，以及识别与锰的获取、运输和储存有关的蛋白质。有关锰代谢的详细情况可能会浮出水面。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研究生提供出色的培训，并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。 公共卫生相关性：自由基介导的损害与200多种人类疾病和衰老有关。这个拟议的项目利用一种简单的、活的、多细胞的、与人类相似的真核生物，来研究潜在的自由基清除剂--锰--如何在细胞内发挥作用。了解锰是如何进入细胞发挥其有益的抗氧化剂样功能的，这与人类健康有关，因为它可以导致新的药物或补充剂，这些药物或补充剂可能会减缓衰老过程，提高生活质量。

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.