The Biological Role of Heme in Human Nutrition

世界上五分之四的人可能缺铁，使营养性缺铁成为最常见的铁缺乏症。 常见的营养失调虽然大量的实验和营养数据支持一个基本的 血红素（铁-原卟啉IX）作为人体营养中铁的生物可利用来源的作用， 血红素吸收和利用目前是未知的。从细胞生物学的角度来看，血红素是 辅基是许多重要生物过程的辅基，在真核生物中，辅基是在细胞中合成的。 线粒体基质因为血红素是一种疏水分子，并且由于其内在的 过氧化物酶活性，我们假设血红素不仅通过脂质双层扩散， 通过特定的细胞内途径运输。我们建议使用的动物模型，小杆线虫 elegans，以确定血红素运输的细胞途径和介导血红素运输的分子， 因为这种动物是唯一已知的不能合成血红素眼的遗传动物模型， 需要血红素来维持代谢过程。自从C.秀丽线虫缺乏制造血红素的能力， 我们有一个干净的遗传背景缺乏内源性血红素，并有能力从外部控制 血红素代谢通量因此，C.线虫是一种专性血红素营养缺陷型， 模型研究膳食血红素吸收以及细胞内血红素运输，螯合 并掺入关键血红素蛋白中。血红素转运的细胞途径将在C. 通过生物化学表征从正向遗传筛选中分离的突变体的表型簇， 测定血红素水平和血红素蛋白活性，评估存活率并跟踪血红素在肝脏中的转运 动物与血红素类似物，并确定血红素分布在超微结构水平上使用电子 显微镜突变基因的分子身份将通过对突变体进行分类来确定， 遗传互补组，使用SNP定位突变基因，通过 RNA干扰和基因拯救，并通过对候选基因进行测序来确定分子损伤。 C.从我们的DNA微阵列中鉴定出的线虫，其特征在于 通过实时PCR和RNA印迹分析定量验证基因的子集， 利用RNA干扰技术对候选基因进行筛选，确定候选基因的时空表达 通过合成转录和翻译报告基因融合体来产生对血红素应答的产物。的 这些研究的结果将为真核生物中血红素的稳态提供新的机制见解， 可能有助于开发针对人类缺铁的基于血红素的营养干预措施， 人类蠕虫感染的潜在药物靶点，加剧缺铁。

Four out of five people in the world may be iron-deficient, making nutritional iron deficiency the most common nutritional disorder. Although considerable experimental and nutritional data support an essential role for heme (iron-protoporphyrin IX) as a bioavailable source of iron in human nutrition, the pathways for heme absorption and utilization are currently unknown. From a cell biological perspective, heme is the prosthetic group for many important biological processes, and in eukaryotes it is synthesized in the mitochondrial matrix. Because heme is a hydrophobic molecule and is cytotoxic due to its intrinsic peroxidase activity, we hypothesize that heme does not merely diffuse through lipid bilayers but is actively transported via specific intracellular pathways. We propose to use the animal model, Caenorhabditis elegans, to identify the cellular pathways for heme transport and the molecules mediating heme trafficking, because this animal is the only known genetic animal model that is unable to synthesize heme eye no^o albeit requiring heme to sustain metabolic processes. Since C. elegans lacks the ability to make heme, it provides us with a clean genetic background devoid of endogenous heme, and the capacity to externally control the metabolic flux of heme. Thus, C. elegans is an obligate heme auxotroph and will be an excellent animal model to study dietary heme absorption as well as intracellular heme trafficking for transport, sequestration and incorporation into critical hemoproteins. The cellular pathways for heme transport will be mapped in C. elegans by characterizing phenoclusters of mutants isolated from forward genetic screens by biochemically assaying for heme levels and hemoprotein activity, evaluating viability and tracking heme transport in live animals with heme analogs, and determining heme distribution at the ultrastructural level using electron microscopy. The molecular identities of the mutated genes will be determined by categorizing mutants in genetic complementation groups, mapping the mutated genes using SNPs, localizing the mutant gene by RNA interference and gene rescue, and by sequencing candidate genes to determine the molecular lesion. The heme regulated genes in C. elegans, identified from our DMA microarrays, will be characterized by quantitatively validating subsets of genes by real-time PCR and RNA blot analysis, determining the function of the candidate genes by RNA interference, and defining the temporal and spatial expression of these gene products in response to heme by synthesizing transcriptional and translational reporter gene fusions. The results from these studies will provide new mechanistic insights into heme homeostasis in eukaryotes and may aid in the development of heme-based nutritional interventions for human iron deficiency, and potential drug targets for human helminthic infections that exacerbate iron deficiency.