Harnessing the Lysosome Machinery to Counter Metal Toxicity

镉是一种有毒金属，对人类健康有重大危害。我们迫切需要发展

Cadmium is a toxic metal and a significant human health hazard. There is an urgent need to develop therapeutic strategies that target the cellular and molecular mechanisms of injury. Our long-term goal is to identify and characterize endogenous mechanisms of cadmium detoxification that can be adapted for therapeutic benefit. Using the powerful C. elegans model system, we discovered that exposure to high dietary zinc stimulates lysosome biogenesis by activating the master transcriptional regulator, TFEB (HLH-30 in C. elegans). Cadmium is similar to zinc, and cadmium exposure causes a transcriptional response similar to zinc excess. We propose the innovative hypothesis that TFEB plays an important role in cadmium detoxification by enhancing lysosome biogenesis, leading to increased metal sequestration, and by enhancing mitochondrial quality. Here we propose to rigorously test this hypothesis by conducting two Specific Aims using a wide range of genetic and cell biology techniques. We will exploit C. elegans as well as the clinically relevant mouse model and explore the use of trehalose as a potential therapeutic that activates TFEB. Aim 1: Characterize the function and regulation of the HLH-30/TFEB signaling axis during cadmium exposure in C. elegans and mice. We will analyze accumulation of cadmium and the role of lysosomes in cadmium resistance in C. elegans. Genetic analysis using gain-of-function and loss-of-function approaches will rigorously determine the function of hlh-30 during cadmium toxicity. The regulation of hlh-30 by cadmium exposure will be determined in C. elegans. To establish the function of TFEB during cadmium exposure in mice, we will generate genetically targeted mice that over-express or lack TFEB specifically in hepatocytes or in renal tubular cells. Liver or renal injury will be measured after cadmium exposure. We will determine how cadmium exposure regulates TFEB signaling in murine hepatocytes and proximal renal tubular cells. We will determine if administration of trehalose, a naturally occurring non-reducing sugar which activates TFEB, will be effective in preventing and treating cadmium-induced hepatotoxicity and nephrotoxicity in mice. Genetic ablation of the enzyme trehalase will be analyzed to determine the efficacy of increased trehalose bioavailability on cadmium-induced toxicity. Aim 2: Determine cellular mechanisms of cadmium toxicity by analyzing mitochondria and lysosome remodeling. We will determine how cadmium and hlh-30/TFEB activity effect mitochondria in C. elegans, mice, and mammalian cells. Measurements will include mitochondrial ROS generation, mass, ultrastructure, and function. These studies will rigorously test the model that mitochondrial damage plays a role in cadmium toxicity and can be ameliorated by HLH-30/ TFEB. We will determine how cadmium and hlh-30 activity effect lysosomal structural remodeling in C. elegans and mammalian cells using state-of-the-art super resolution microscopy. Successful completion of these experiments will have a high impact by elucidating the role of TFEB in cadmium resistance and the viability of trehalose as an approach to ameliorate cadmium toxicity.