Illuminating Heme Trafficking and Signaling Pathways in Health and Disease

项目总结 这项提议的长期目标是阐明调节血红素的分子和机制。 在健康和疾病方面的贩运和信号。血红素是一种重要的但具有潜在细胞毒性的金属辅因子， 信号分子，铁的营养来源。所有依赖于血红素的过程都要求血红素是安全的 从摄取或合成部位运输和贩运到依赖于血红素或受调控的蛋白质 遍及整个细胞。然而，介导血红素动员的分子和机制 人们对贩卖和发送信号知之甚少。此外，血红素运输的时空动态是 未知。这在很大程度上是因为缺乏工具来对与之相关的不稳定的血红素库进行成像 生物分布。雷迪实验室最近克服了这些技术障碍，通过产生基因 编码比率荧光血红素传感器和表征血红素的血液蛋白质组学方法 贩运和信号动态。这项提议将重点放在将分子遗传学、生化、 生物物理、细胞生物学和化学生物学方法以阐明：A.血红素的运输和 贩运；B.基于血红素的信号转导；以及通过各种模式真核生物获得C.heme 细胞系，如酿酒酵母和哺乳动物细胞系，以及病原体，如分枝杆菌 肺结核和白色念珠菌。为了探测细胞内的血红素运输，基因编码的血红素传感器 将用于基因缺失和过度表达筛查，以确定血红素分布和 贩卖人口。为了探索血红素信号，将部署血液蛋白质组学方法和血红素螯合剂来 确定血红素信号是如何启动、传递和整合以控制新陈代谢和生理的。至 通过人类病原体、血红素传感器和血液蛋白质组学方法探测血红素的获取 综合鉴定结核分枝杆菌对血红素的吸收和利用机制 白色念珠菌。总之，拟议的研究将全面解决血红素的作用机制。 在与健康和疾病有关的各种情况下进行贩运和发送信号。

PROJECT SUMMARY The long-term objectives of this proposal are to elucidate the molecules and mechanisms that mediate heme trafficking and signaling in health and disease. Heme is an essential but potentially cytotoxic metallocofactor, signaling molecule, and nutritional source of iron. All heme-dependent processes require that heme is safely transported and trafficked from sites of uptake or synthesis to heme dependent or regulated proteins located throughout the cell. However, the molecules and mechanisms that mediate the mobilization of heme for trafficking and signaling are poorly understood. Further, the spatio-temporal dynamics of heme trafficking are unknown. This is in large part due to the lack of tools available to image labile heme pools relevant to it biodistribution. The Reddi lab has recently overcome these technological barriers by generating genetically encoded ratiometric fluorescent heme sensors and hemoproteomics approaches to characterize heme trafficking and signaling dynamics. This proposal will focus on integrating molecular genetic, biochemical, biophysical, cell biological, and chemical biology approaches towards elucidating: a. heme transport and trafficking; b. heme-based signal transduction; and c. heme acquisition across a variety of model eukaryotic cell lines, e.g. Saccharomyces cerevisiae and mammalian cell lines, and pathogens, e.g. Mycobacterium tuberculosis and Candida albicans. To probe intracellular heme trafficking, genetically encoded heme sensors will be used in gene deletion and over-expression screens to identify novel pathways for heme distribution and trafficking. To probe heme signaling, hemoproteomics approaches and heme chelators will be deployed to identify how heme signals are initiated, transmitted, and integrated to control metabolism and physiology. To probe heme acquisition by human pathogens, heme sensors and hemoproteomics approaches will be integrated to identify the mechanisms of heme uptake and utilization by Mycobacterium tuberculosis and Candida albicans. Altogether, the proposed research will comprehensively address the mechanisms of heme trafficking and signaling in various contexts relevant to health and disease.