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，对铁代谢的见解迅速增长，鉴定了参与铁吸收和运输的蛋白质，以及对铁在基因表达中的作用的认识。最近鉴定出的蛋白质是二价金属离子转运蛋白I（DMT1），该转运蛋白I（DMT1）在粉红细胞中的十二指肠肠肠细胞和铁转运在铁吸收中起作用。 DMT1 mRNA在每个组织中都发现，但是其在不参与铁代谢的组织中的作用尚不清楚。对DMT1 mRNA的评估揭示了分子的5'和3'端的替代剪接，导致至少4种同工型，似乎在各种组织中分布不同。在啮齿动物中，DMT1基因突变会导致低色素/微细胞贫血和铁缺乏症，但是，在患有低染色/微细胞贫血和铁超载的女性中发现了唯一已知的人DMT1突变。人类突变预测了保守的氨基酸取代，但其主要作用是跳过外显子12。在正常受试者中，大约10％的网状细胞DMT1 mRNA是外显子12跳过的转录本，表明该突变夸大了“正常”过程。促红细胞生成素或缺氧的上型浓度在体外纠正了患者红细胞细胞的表型。这种校正是在分子水平上发生的方式，这是人类患者铁超载的细节。铁如何调节DMT1 mRNA水平以及缺氧，红细胞生成素和细胞因子发挥的作用的细节尚不清楚。外显子跳过的功能及其调节也有待探讨。该建议通过检查控制DMT1 mRNA的因素和各种组织中的蛋白质水平来解决这些问题。铁状态，缺氧，红细胞生成素和细胞因子水平对转录，消息稳定性和替代剪接的影响将在人类细胞系和原发性红细胞细胞培养物中进行评估。一旦确定了这些因素，Exon 12跳过的组织特异性，Exon 12与特定转录物的关联以及其按影响DMT1 mRNA水平的因素进行调节。对各种组织中DMT1调控的欣赏将有助于进一步阐明其在铁代谢中涉及的组织中的功能，但也没有。这些知识不仅在治疗贫血患者方面是无价的，而且在治疗铁超负荷的患者或铁超载特定组织有助于疾病的患者方面也是无价的。