Inter-organ signaling in heme metabolism and organismal homeostasis.

本提案的长期目标是确定机体血红素稳态的细胞和分子决定因素。血红素是一种含铁的有机环，是多种生物功能的重要辅助因子，也是人类饮食中生物可利用铁的主要来源。作为一种疏水和细胞毒性辅助因子，血红素必须通过特定的细胞内和细胞间途径以高度控制的方式通过膜进行运输。然而，负责血红素运输的基因和途径仍然知之甚少。目前的范式表明，细胞对血红素的需求是通过细胞内部调节和合成自身血红素的能力来实现的。我们的范式转换假设是，细胞血红素水平不仅通过内部血红素合成（细胞自主）维持，而且还通过远端定位的蛋白质来维持，这些蛋白质向器官间血红素运输网络发出系统性血红素需求的信号（细胞非自主）。尽管新出现的证据支持哺乳动物系统细胞-非自主血红素通信系统的存在，但这一概念仍未被探索。秀丽隐杆线虫是一种独特的动物模型，可以确定是否存在这样的系统信号通路，因为秀丽隐杆线虫允许系统血红素稳态被营养和遗传操纵，并且它们的光学透明性允许在亚细胞分辨率下监测组织之间的血红素信号。在上一个资助期，我们确定了肠和神经元之间血红素状态的一种新的双向信号通路。我们发现，远端器官具有专门的机制，通过HRG-7（组织蛋白酶E）、DBL-1 （BMP5）、SMA-9 （SHN）和HRG-1 （SLC48A1）与关键的血红素储存和运输器官-肠进行沟通和协调其血红素状态。在本提案中，我们将验证器官间血红素通信由HRG-7依赖和HRG-7独立途径协调的假设。这一假设得到了来自蛋白质组学、全基因组RNAi、正向遗传和RNAseq研究的令人兴奋的新初步数据的支持。我们将(a)阐明分泌egf结构域蛋白直接调节hrg -7依赖性信号通路的分子机制；(b)评估在调节hrg -7不依赖的组织间血红素信号通路中跨膜钾离子通道的分子需求；(c)揭示衔接蛋白复合物协调肠道血红素运输和肠外血红素分配以调节全身血红素信号传导的机制。我们的目标是在完整动物的背景下，在组织和亚细胞分辨率上深入了解血红素依赖性信号，因为人类体内95%的铁配额都在血红素蛋白中。

The long-term goals of this proposal are to define the cellular and molecular determinants of organismal heme homeostasis. Heme, an iron-containing organic ring, functions as a vital cofactor responsible for diverse biological functions, and is the major source of bioavailable iron in the human diet. As a hydrophobic and cytotoxic cofactor, heme must be transported in a highly controlled manner through membranes via specific intra- and inter-cellular pathways. However, the genes and pathways responsible for heme trafficking remain poorly understood. The current paradigm states that cellular requirements for heme are fulfilled by the cell’s internal capacity to regulate and synthesize its own heme. Our paradigm-shifting hypothesis is that cellular heme levels are not only maintained by internal heme synthesis (cell-autonomous), but also by distally located proteins which signal systemic heme requirements to an inter-organ heme trafficking network (cell- nonautonomous). Although emerging evidence support the existence of a systemic cell-nonautonomous heme communication system in mammals, this concept has remained unexplored. Caenorhabditis elegans is a unique animal model to determine if such systemic signaling pathways exist as C. elegans allows systemic heme homeostasis to be manipulated nutritionally and genetically, and their optical transparency allows for in vivo monitoring of heme signals between tissues at subcellular resolution. In the last funding period, we identified a novel bidirectional signaling pathway of heme status between the intestine and neuron. We showed that distal organs have dedicated mechanisms to communicate and coordinate their heme status with the crucial heme storage and transport organ – the intestine via HRG-7 (cathepsin E), DBL-1 (BMP5), SMA-9 (SHN) and HRG-1 (SLC48A1). In this proposal, we will test the hypothesis that inter-organ heme communication is coordinated by both, HRG-7 dependent and HRG-7 independent pathways. This hypothesis is supported by exciting new preliminary data from proteomic, genome-wide RNAi, forward genetic, and RNAseq studies. We will (a) elucidate the molecular mechanisms for a secreted EGF-domain protein in directly regulating HRG-7-dependent signaling pathway; (b) assess the molecular requirement for a transmembrane potassium/ion channel in regulating HRG-7-independent inter-tissue heme signaling pathway; and (c) uncover the mechanisms for adaptor protein complexes in coordinating intestinal heme transport with extra-intestinal heme allocation to regulate systemic heme signaling. Our goals are to acquire a deep understanding of heme-dependent signaling at a tissue and subcellular resolution within the context of an intact animal, as 95% of the total body iron quota in humans is within heme proteins.