MECHANISTIC INSIGHTS INTO CADMIUM DETOXIFICATION

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 中心，不一定是研究者的机构。 非生理金属的去铁化和营养但有毒金属的稳态获得是基本的生物过程。镉是一种毒性很强的环境污染物，可引起多种人类疾病，包括肾脏疾病、癌症和内分泌紊乱。氧化性细胞损伤、营养金属稳态的扰动、DNA修复的抑制和雌激素活性与镉毒性有关。然而，镉在真核生物中的解毒机制，特别是镉排泄系统，在很大程度上是未知的。该项目的长期目标是表征镉解毒的分子机制，并利用这些知识减少人类接触镉。在寻找酿酒酵母中与重金属抗性相关的基因的过程中，我们鉴定了一个P型ATP酶。从细菌到人类的所有生物体都依赖于这个转运蛋白家族来维持各种离子的跨膜梯度。我们的数据强烈表明，这P型ATP酶是一个镉选择性出口商。此外，当细胞在含镉培养基中生长时，这种转运蛋白的表达水平通过镉介导的活性周转抑制而迅速上调。该应用程序的重点是表征的功能，作用机制和调节的镉转运蛋白。中心假设是，这种P型ATP酶是第一个镉特异性外排泵，其在结构、底物特异性和调节模式上是独特的。这一假设将使用生物化学、细胞生物学和遗传学方法进行检验。首先，将阐明P型ATP酶的金属特异性。本研究将主要集中在体内金属抗性和积累测定和体外ATP酶测定。其次，结合酵母遗传学，细胞生物学和化学的多学科方法将确定参与这种P型ATP酶的镉依赖性翻译后控制的独特模式的调控元件。拟议的研究将揭示一种新的镉解毒机制，由酵母中的P型ATP酶介导，酵母是一种模式真核生物，并最终提高我们对抗人类镉相关疾病的能力。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Detoxification of non-physiological metals and homeostatic acquisition of nutritional yet toxic metals are fundamental biological processes. Cadmium is a highly toxic environmental contaminant, which causes a number of human disorders, including kidney disease, cancer, and endocrine disruption. Oxidative cellular damage, perturbation of nutritional metal homeostasis, inhibition of DNA repair, and estrogenic activities are implicated with cadmium toxicity. However, the mechanisms of cadmium detoxification in eukaryotes, especially cadmium excretion systems, are largely unknown. The long-term goals of this project are the characterization of molecular mechanisms of cadmium detoxification and employing this knowledge to reduce cadmium exposure to humans. During the search for genes involved in heavy metal resistance in yeast Saccharomyces cerevisiae, we have identified a P-type ATPase. All organisms ranging from bacteria to humans rely on this family of transporters for maintaining a trans-membrane gradient of various ions. Our data strongly suggest that this P-type ATPase is a cadmium selective exporter. Moreover, when cells grow in cadmium-containing media, the expression levels of this transporter are rapidly up regulated through cadmium-mediated inhibition of active turnover. This application focuses on characterization of the function, mechanisms of action and regulation of this cadmium transporter. The central hypothesis is that this P-type ATPase is the first cadmium-specific efflux pump that is unique in structure, substrate specificity and mode of regulation. This hypothesis will be tested using biochemical, cell biological and genetic approaches. First, metal specificity of the P-type ATPase will be elucidated. This study will largely focus on in vivo metal resistant and accumulation assays and in vitro ATPase assays. Second, a multi-disciplinary approach combining yeast genetics, cell biology and chemistry will identify regulatory elements involved in the unique mode of cadmium-dependent post-translational control of this P-type ATPase. The proposed studies will reveal a novel cadmium detoxification mechanism mediated by a P-type ATPase in yeast, a model eukaryote, and ultimately advance our ability to combat cadmium related disorders in humans.