DMT1:Tissue specific expression & alternative splicing

DMT1：组织特异性表达

• 批准号: 7031607
• 负责人: Martha P. Mims
• 金额: $ 14.65万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7031607
• 关键词: RNA splicing; biological transport; cell line; clinical research; cytokine; erythropoietin; gene expression profiling; human tissue; hypoxia; iron metabolism; iron poisoning; messenger RNA; transcription factor; transport proteins

DESCRIPTION (provided by applicant): Insight into iron metabolism has grown rapidly in recent years with identification of proteins involved in iron absorption and trafficking, and recognition of the role of iron in gene expression. One recently identified protein is divalent metal ion transporter I (DMT1) which functions in iron absorption by the duodenal enterocyte and iron transport in the erythroblast. DMT1 mRNA is found in every tissue, but its role in tissues not involved in iron metabolism is unclear. Evaluation of DMT1 mRNA reveals alternative splicing at both the 5' and 3' ends of the molecule leading to at least 4 isoforms, which seem to be distributed differently in various tissues. In rodents, DMT1 gene mutation leads to hypochromic/microcytic anemia and iron deficiency, however, the only known human DMT1 mutation was found in a woman with hypochromic/microcytic anemia and iron overload. The human mutation predicts a conservative amino acid substitution, but its predominant effect is skipping of exon 12. In normal subjects, approximately 10% of reticulocyte DMT1 mRNA is exon 12 skipped transcript suggesting that the mutation exaggerates a "normal" process. Supratherapeutic concentrations of erythropoietin or hypoxia correct the phenotype of the patient's erythroid cells in vitro. How this correction occurs at the molecular level is unknown as are the specifics of why the human patient is iron overloaded. The details of how iron regulates DMT1 mRNA levels, and the roles that hypoxia, erythropoietin, and cytokines play are unclear. The function of exon 12 skipping, and its regulation also remain to be explored. This proposal addresses these issues by examining factors, which control DMT1 mRNA, and protein levels in various tissues. The effects of iron status, hypoxia, erythropoietin and cytokine levels on transcription, message stability and alternative splicing will be evaluated in human cell lines and primary erythroid cell cultures. Once these factors have been identified, the tissue specificity of exon 12 skipping, the association of exon 12 skipping with a specific transcript, and its modulation by factors which affect DMT1 mRNA levels will be examined. Appreciation of DMT1 regulation in various tissues will help further unravel its function(s) in tissues involved in iron metabolism, but also in tissues, which are not. This knowledge will be invaluable not only in treating anemic patients, but also in treating patients who are iron overloaded or in whom iron overload of specific tissues contributes to disease.

描述（由申请人提供）：近年来，随着参与铁吸收和运输的蛋白质的鉴定以及铁在基因表达中的作用的认识，对铁代谢的了解迅速增长。最近发现的一种蛋白质是二价金属离子转运蛋白I (DMT1)，它在十二指肠肠细胞铁吸收和红母细胞铁运输中起作用。DMT1 mRNA存在于所有组织中，但其在非铁代谢组织中的作用尚不清楚。对DMT1 mRNA的评估显示，在分子的5‘和3’端选择性剪接导致至少4种异构体，这些异构体似乎在不同组织中分布不同。在啮齿动物中，DMT1基因突变导致低色素/小细胞贫血和缺铁，然而，唯一已知的人类DMT1突变是在一名患有低色素/小细胞贫血和铁超载的女性中发现的。人类突变预测保守的氨基酸取代，但其主要影响是外显子12的跳过。在正常受试者中，大约10%的网织红细胞DMT1 mRNA是12外显子跳过的转录本，这表明该突变夸大了“正常”过程。超治疗浓度的促红细胞生成素或缺氧可在体外纠正患者红细胞的表型。这种纠正是如何在分子水平上发生的尚不清楚，也不清楚人类患者铁超载的具体原因。铁如何调节DMT1 mRNA水平的细节，以及缺氧、促红细胞生成素和细胞因子在其中的作用尚不清楚。外显子12跳变的功能及其调控也有待探讨。本提案通过检查控制各种组织中DMT1 mRNA和蛋白质水平的因素来解决这些问题。铁状态、缺氧、促红细胞生成素和细胞因子水平对转录、信息稳定性和选择性剪接的影响将在人类细胞系和原代红细胞培养中进行评估。一旦确定了这些因素，将研究外显子12跳变的组织特异性，外显子12跳变与特定转录物的关联，以及影响DMT1 mRNA水平的因素对其的调节。了解DMT1在各种组织中的调节将有助于进一步揭示其在参与铁代谢的组织中的功能，以及在不参与铁代谢的组织中的功能。这一知识不仅对治疗贫血患者，而且对治疗铁超载或特定组织铁超载导致疾病的患者都是无价的。