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。赖氨酰氧化酶 (LO) 是一种铜 (Cu) 依赖性酶，可氧化肽基赖氨酸基质中的残留物，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。该酶已被鉴定为肿瘤抑制因子，例如，抑制原癌基因ras的转化活性。因此，LO 作为细胞内和细胞外效应器在人体生理和病理中起着至关重要的作用。人类长期接触镉 (Cd)，一种重金属，无论是来自职业污染还是来自香烟烟雾，都会引起肺气肿和肺癌。然而，镉引起的肺部病理学机制仍知之甚少。 LO 作为金属酶容易受到细胞金属稳态变化的影响。该实验室之前的研究研究大鼠肺成纤维细胞 (RFL6) 从镉敏感到镉抗性的表型变化 Cd 在 mRNA、蛋白质和催化水平上下调 LO。持续研究进一步表明 RFL6 细胞对 Cd 的反应表现出 LO 转录起始的抑制和 LO mRNA 的增强衰变共同导致稳态 LO mRNA 水平下降。这些发现导致 LO 基因的转录控制和调节是 Cd 损伤的关键目标的假设 Cd 对 LO 基因转录的沉默是肺部疾病的关键分子基础。该项目的总体目标是拟议的研究是通过实现以下具体目标来检验这一假设：1）确定机制通过检查 RNA 聚合酶 II 的 Cd 调节，在转录水平上对 LO 基因进行 Cd 沉默定向 LO 前 mRNA 合成和加工，以及核心调控的 LO 启动子激活发起人； 2) 通过检查确定 LO 基因在转录水平上的 Cd 沉默机制 Cd 对 LO 启动子激活的调节受金属和氧化还原敏感转录因子的调节它们的同源顺式元件，并确定 LO 启动子甲基化，为 Cd 表观遗传提供证据 LO DNA 损伤； 3) 确定转录后水平上LO基因Cd沉默的机制通过检查 Cd 对 LO mRNA 5'-加帽和 3'-聚腺苷酸化状态的影响，并评估 Cd 3'-非翻译区中敏感的、LO mRNA 稳定性相关的顺式元件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 改变的 LO 表达对底物启动子激活和细胞的影响来建立模型细胞模型中的转化并评估主要 LO 转录和转录后机制以及肺气肿中 LO 基因启动子的异常甲基化以及长期服用镉的大鼠的致癌肺部。拟议研究的结果预计将增强我们对 Cd 沉默 LO 基因机制的理解，为制定镉相关肺部疾病的预防和治疗策略。