Lung Lysyl Oxidase Regulation by Metal Ion Homeostasis

金属离子稳态对肺赖氨酰氧化酶的调节

基本信息

批准号：
8449745
负责人：
Wande Li
金额：
$ 35.12万
依托单位：
BOSTON UNIVERSITY MEDICAL CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
2003
资助国家：
美国
起止时间：
2003-06-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8449745
关键词：
5&apos Flanking Region Affect Animal Model Atherosclerosis Binding Proteins Biological Cadmium Cell Nucleus Cell model Cells Chronic Cigarette smoke-induced emphysema Collagen Copper DNA Disease Dose Down-Regulation Elastin Elements Enzymes Epigenetic Process Exhibits Extracellular Matrix Fibroblasts Fibrosis Gene Silencing Genes Genetic Transcription Goals Half-Life Health Heavy Metals Histone H1 Histone H1(s)Homeostasis Human Indium Industry Ions Lung Lung diseases Lysine Malignant Neoplasms Malignant neoplasm of lung Messenger RNA Metals Methylation Modeling Molecular Morphogenesis Occupational Organ Outcome Oxidation-Reduction Pathogenesis Pathology Phenotype Physiology Play Polyadenylation Process Prophylactic treatment Protein-Lysine 6-Oxidase Proteins Proto-Oncogenes Pulmonary Emphysema RNA Polymerase II Rattus Regulation Research Resistance Role Source Testing Therapeutic Tobacco Transcript Transcription Initiation Transcriptional Regulation Tumor Suppressor Proteins base carcinogenesis catalyst cell transformation cigarette smoking exposed human population extracellular lung basal segment lung tumorigenesis mRNA Decay mRNA Precursor mRNA Stability metalloenzyme promoter response tissue repair transcription factor treatment strategy

项目摘要

PROJECT ABSTRACT. Lysyl oxidase (LO), a copper-(Cu) dependent enzyme, oxidizes peptidyl lysine residues in substrates, e.g., collagen, elastin and histone H1, essential for organization and stabilization of the extracellular matrix (ECM) and the cell nucleus. This enzyme has been identified as a tumor suppressor, for example, inhibiting transforming activity of ras, a proto oncogene. Thus, LO as an intra- and extracellular effector plays a critical role in human physiology and pathology. Chronic exposure of humans to cadmium (Cd), a heavy metal, either from occupational contamination or from cigarette smoke, induces emphysema and lung cancers. However, the mechanisms for Cd-elicited lung pathology remain poorly understood. LO as a metalloenzyme is susceptible to changes in cellular metal homeostasis. Previous studies by this lab investigating the phenotype change from Cd sensitive to Cd resistant of rat lung fibroblasts (RFL6) illustrated downregulation of LO by Cd at mRNA, protein and catalytic levels. Continuing studies further indicated that RFL6 cells in response to Cd displayed inhibition of LO transcription initiation and enhancement of LO mRNA decay both collectively contributing to decreased levels of steady-state LO mRNAs. These findings have led to a hypothesis that transcriptional control and regulation of the LO gene are critical targets for Cd insult and silencing of LO gene transcription by Cd is a key molecular basis for lung diseases. The overall goal of the proposed research is to test this hypothesis by achieving following specific aims: 1) to identify mechanisms for Cd silencing of the LO gene at the transcriptional level by examining Cd modulation of RNA polymerase II- directed LO pre-mRNA synthesis and processing, and of the LO promoter activation regulated by the core promoter; 2) to identify mechanisms for Cd silencing of the LO gene at the transcriptional level by examining Cd modulation of the LO promoter activation regulated by metal and redox-sensitive transcription factors and their cognate cis-elements, and determining LO promoter methylation to provide evidence for Cd epigenetic damage to LO DNA; 3) to identify mechanisms for Cd silencing of the LO gene at the posttranscriptional level by examining Cd effects on the 5'-capping and 3'-polyadenylation status of LO mRNA, and assessing Cd sensitive, LO mRNA stability-related cis-elements in the 3'-untranslation region and their corresponding binding proteins; and 4) to investigate biological consequences of Cd silencing of the LO gene in cell and animal models by examining effects of altered LO expression by Cd on substrate promoter activation and cell transformation in the cell model and assessing the active status of the major LO transcriptional and posttranscriptional machineries as well as aberrant methylation of the LO gene promoter in emphysematous and carcinogenic lungs of rats receiving Cd by chronic administration. The outcomes of the proposed research are expected to enhance our understanding of mechanisms of LO gene silence by Cd providing the basis for developing prophylaxis and treatment strategies for Cd-related lung diseases.

项目摘要。赖氨酸氧化酶（LO），一种铜（Cu）依赖性酶，氧化肽基赖氨酸底物中的残基，例如胶原蛋白，弹性蛋白和组蛋白H1，对于组织和稳定至关重要细胞外基质（ECM）和细胞核。该酶已被确定为肿瘤抑制剂，因为例如，抑制原始致癌基因Ras的转化活性。因此，LO作为细胞内和细胞外效应子在人类生理和病理学中起关键作用。人类长期暴露于镉（CD），一种重金属，无论是从职业污染还是来自香烟烟雾，都会引起肺气肿和肺癌。然而，CD引起的肺病理学的机制仍然鲜为人知。 lo作为一个金属酶容易受到细胞金属稳态变化的影响。该实验室的先前研究研究表型从CD敏感到抗CD的大鼠肺成纤维细胞（RFL6）的变化所示在mRNA，蛋白质和催化水平下通过CD下调LO。继续研究进一步表明响应CD的RFL6细胞显示出对LO转录启动和LO mRNA的增强的抑制衰减均集体促进稳态LO mRNA水平降低。这些发现导致了一个假说，即转录控制和调节基因是CD损伤和的关键目标通过CD沉默LO基因转录是肺部疾病的关键分子基础。总体目标拟议的研究是通过实现以下特定目的来检验这一假设：1）确定机制通过检查RNA聚合酶II-的CD调节，用于转录水平的LO基因沉默定向LO前MRNA合成和加工，以及由核心调节的LO启动子激活发起人； 2）通过检查转录水平的LO基因的CD沉默机制由金属和氧化还原敏感的转录因子调节的LO启动子激活的CD调节以及它们的同源顺式元素，并确定LO启动子甲基化以提供CD表观遗传学的证据对LO DNA的损害； 3）确定在转录后水平上LO基因的CD沉默的机制通过检查CD对LO mRNA的5'限制和3--聚乙酰化状态的影响，并评估CD 3'-非翻译区域中敏感的，LO mRNA稳定性相关的顺式元素及其相应的结合蛋白质； 4）研究细胞和动物中LO基因CD沉默的生物学后果通过检查CD改变LO表达对底物启动子激活和细胞的影响模型细胞模型中的转换并评估主要LO转录的活动状态和转录后机器以及LO基因启动子的异常甲基化慢性给药接受CD的大鼠的致癌肺。拟议研究的结果期望通过CD为LO基因沉默的机制增强我们的理解开发针对CD相关肺部疾病的预防和治疗策略。