The Transport of Nutritional Heme in Animal Development

 描述(由申请人提供)：这项提案的长期目标是定义人类营养中的血红素稳态的细胞和分子决定因素。血红素是肠细胞和巨噬细胞生物可利用铁的重要来源，在肠细胞中，血红素铁被吸收，在巨噬细胞中，血红素铁被循环利用。在人体肠道中，膳食中的血红素比无机铁更容易被吸收，也是西方饮食中体内大部分铁的来源。此外，超过70%的体内铁以血红素的形式存在于血红蛋白中。血红素中的铁通过巨噬细胞的吞噬小体中衰老的红细胞的吞噬来循环利用。到目前为止，负责血红素在人类肠道和网状内皮系统的巨噬细胞中运输的基因和途径仍不清楚。作为一种疏水性和细胞毒性的辅因子，血红素必须通过特定的细胞内和细胞间途径通过膜以高度受控的方式运输。在上一个供资期间之前，纽约州的血红素进口商的身份不详。我们认为线虫是鉴定血红素转运途径的一个很好的动物模型，因为它能合成大量具有人类同源物的血红素蛋白，但不能从头合成血红素。通过利用前一个资助周期中线虫的血红素营养缺陷症，我们成功地鉴定了第一个真核细胞中的血红素进口/转运体HRG1及其相应的人类同源物，这一发现为血红素在动物中的运输和贩运提供了第一个初步的框架。HRG1对巨噬细胞铁稳态是必不可少的，在衰老的红细胞吞噬过程中，它将血红素从吞噬小体运送到细胞质--这一过程被称为红细胞吞噬作用(细胞代谢2013)。重要的是，我们的研究表明，人类的一个HRG1变种在血红素运输方面存在缺陷。这项建议中的研究旨在阐明HRG1在分子、细胞和生物水平上转运血红素的确切机制。我们试图测试这一假设，即HRG1是难以捉摸的人类肠道血红素转运体，是基本的血红素运输网络的一部分。我们将利用(A)细胞生物学方法来建立HRG1作为肠道血红素转运体；(B)通过遗传学方法来阐明HRG1变异在人类铁代谢紊乱中的分子后果；以及(C)通过生物化学方法来发现HRG1的其他上游和下游相互作用因素，包括一个功能性的血红素运输网络。我们的目标是全面了解在哺乳动物中调节血红素运输的途径，到目前为止，这些途径仍然知之甚少。

 DESCRIPTION (provided by applicant): The long-term goals of this proposal are to define the cellular and molecular determinants of heme homeostasis in human nutrition. Heme is a significant source of bioavailable iron for enterocytes where heme-iron is absorbed and for macrophages where heme-iron is recycled. In the human intestine, dietary heme is more easily absorbed than inorganic iron and is the source for the majority of body iron from a western diet. Moreover, over 70% of the total body iron is present as heme in hemoglobin. Iron from heme is recycled by phagocytosis of senescent red blood cells in the phagolysosome of macrophages. As of yet, the genes and pathways responsible for heme transport in the human intestine and macrophages of the reticuloendothelial system remain poorly defined. As a hydrophobic and cytotoxic cofactor, heme must be transported in a highly controlled manner through membranes via specific intra- and inter-cellular pathways. Prior to the last funding period, the identity of ny heme importer was unknown. We established that the roundworm Caenorhabditis elegans is an excellent animal model to identify heme transport pathways because it synthesizes a large number of hemoproteins with human homologs but does not synthesize heme de novo. By exploiting the heme auxotrophy of C. elegans in the previous grant cycle, we successfully identified the first eukaryotic heme importer/transporter, HRG1, and its corresponding human homolog, a discovery which provided the first preliminary framework for heme transport and trafficking in animals. HRG1 is essential for macrophage iron homeostasis and transports heme from the phagolysosome to the cytoplasm during phagocytosis of senescent RBCs - a process called erythrophagocytosis (Cell Metabolism 2013). Importantly, our studies reveal that a HRG1 variant in humans is defective in heme transport. The studies in this proposal are designed to elucidate the precise mechanisms of heme transport by HRG1 at the molecular, cellular, and organismal levels. We seek to test the hypothesis that HRG1 is the elusive human intestinal heme transporter and part of an essential heme trafficking network. We will utilize (a) a cell biological approach to establish HRG1 as the intestinal heme transporter; (b) a genetic approach to elucidate the molecular consequence of HRG1 variants in human iron metabolism disorders; and (c) a biochemical approach to uncover additional upstream and downstream interactors of HRG1 comprising a functional heme trafficking network. Our goal is to obtain a comprehensive understanding of the pathways which mediate heme transport in mammals that have, heretofore, remained poorly understood.