DMT1:Tissue specific expression & alternative splicing

DMT1：组织特异性表达

• 批准号: 6858234
• 负责人: Martha P. Mims
• 金额: $ 15万
• 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 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6858234
• 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，随着对参与铁吸收和运输的蛋白质的鉴定以及对铁在基因表达中的作用的认识，对铁代谢的了解迅速增长。一种最近鉴定的蛋白质是二价金属离子转运蛋白I（DMT 1），其在十二指肠上皮细胞的铁吸收和成红细胞中的铁转运中起作用。DMT 1 mRNA存在于每个组织中，但其在不参与铁代谢的组织中的作用尚不清楚。DMT 1 mRNA的评价揭示了在分子的5'和3'端的选择性剪接，导致至少4种亚型，其似乎在各种组织中分布不同。在啮齿类动物中，DMT 1基因突变导致低色素/小红细胞性贫血和铁缺乏，然而，唯一已知的人类DMT 1突变发现于一名患有低色素/小红细胞性贫血和铁过载的女性。人类突变预测了保守的氨基酸取代，但其主要影响是外显子12的跳跃。在正常受试者中，大约10%的网织红细胞DMT 1 mRNA是外显子12跳跃的转录本，这表明突变夸大了“正常”过程。超治疗浓度的促红细胞生成素或缺氧在体外纠正患者红系细胞的表型。这种校正如何在分子水平上发生尚不清楚，也不清楚为什么人类患者铁超载。铁如何调节DMT 1 mRNA水平的细节，以及缺氧，促红细胞生成素和细胞因子发挥的作用尚不清楚。外显子12跳跃的功能及其调控也有待进一步研究。该提案通过检查控制各种组织中DMT 1 mRNA和蛋白质水平的因素来解决这些问题。将在人细胞系和原代红细胞系细胞培养物中评价铁状态、缺氧、促红细胞生成素和细胞因子水平对转录、信息稳定性和可变剪接的影响。一旦确定了这些因素，将检查外显子12跳跃的组织特异性、外显子12跳跃与特定转录物的关联以及影响DMT 1 mRNA水平的因素对其的调节。对DMT 1在各种组织中的调节的认识将有助于进一步揭示其在参与铁代谢的组织中的功能，但也有助于了解其在不参与铁代谢的组织中的功能。这些知识不仅在治疗贫血患者方面，而且在治疗铁过载或特定组织铁过载导致疾病的患者方面都是非常宝贵的。