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：
7886756
负责人：
Wande Li
金额：
$ 36.2万
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 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7886756
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 Left 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 Time Tobacco Transcript Transcription Initiation Transcriptional Regulation Tumor Suppressor Proteins Wound Healing base carcinogenesis catalyst cell transformation cigarette smoking exposed human population extracellular lung basal segment lung tumorigenesis mRNA Decay mRNA Precursor mRNA Stability metalloenzyme promoter public health relevance response transcription factor treatment strategy

项目摘要

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: Lysyl oxidase (LO) is a key copper (Cu)-dependent enzyme existing in the extracellular matrix (ECM) and the cell nucleus critical for organ morphogenesis, tissue repair and anti- tumorigenesis of the lung. Previous and preliminary studies by this lab have indicated that cadmium (Cd), a toxic and carcinogenic heavy metal, induced LO gene silencing in rat lung fibroblasts. The proposed research extends previous findings and aims at investigating molecular mechanisms for LO gene silence by Cd in cell and animal models providing the basis for developing protective and therapeutic strategies for Cd-related lung diseases such as emphysema and cancers.

描述（由申请人提供）：赖氨酰氧化酶 (LO) 是一种铜 (Cu) 依赖性酶，可氧化底物中的肽基赖氨酸残基，例如胶原蛋白、弹性蛋白和组蛋白 H1，这对于细胞外基质 (ECM) 的组织和稳定至关重要和细胞核。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类生理学和病理学中发挥着至关重要的作用。人类因职业污染或香烟烟雾而长期接触镉（Cd）这种重金属，会诱发肺气肿和肺癌。然而，镉引起的肺部病理学机制仍知之甚少。 LO 作为一种金属酶，容易受到细胞金属稳态变化的影响。该实验室之前的研究调查了大鼠肺成纤维细胞（RFL6）从镉敏感到镉抗性的表型变化，结果表明镉在 mRNA、蛋白质和催化水平上下调了 LO。持续的研究进一步表明，RFL6 细胞对 Cd 的反应表现出对 LO 转录起始的抑制和 LO mRNA 衰减的增强，两者共同导致稳态 LO mRNA 水平的降低。这些发现提出了这样一个假设：LO 基因的转录控制和调节是 Cd 损伤的关键目标，而 Cd 对 LO 基因转录的沉默是肺部疾病的关键分子基础。拟议研究的总体目标是通过实现以下具体目标来检验这一假设：1）通过检查 RNA 聚合酶 II 指导的 LO 前 mRNA 合成的 Cd 调节，确定转录水平上 LO 基因的 Cd 沉默机制。核心启动子调控的LO启动子激活的加工和激活； 2) 通过检查由金属和氧化还原敏感转录因子及其同源顺式元件调节的LO启动子激活的Cd调节，并确定LO启动子甲基化，从而确定转录水平上LO基因Cd沉默的机制，并确定LO启动子甲基化，为以下方面提供证据： Cd 对 LO DNA 的表观遗传损伤； 3) 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 敏感性、LO mRNA 稳定性相关顺式元件，确定转录后水平的 LO 基因 Cd 沉默机制。 3'-非翻译区及其相应的结合蛋白； 4) 通过检查 Cd 改变 LO 表达对细胞模型中底物启动子激活和细胞转化的影响，并评估主要 LO 转录和转录的活性状态，研究细胞和动物模型中 LO 基因 Cd 沉默的生物学后果。长期接受镉治疗的大鼠肺气肿和致癌肺中转录后机制以及 LO 基因启动子的异常甲基化。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 Cd 相关肺部疾病的预防和治疗策略奠定基础。公共卫生相关性：赖氨酰氧化酶 (LO) 是一种关键的铜 (Cu) 依赖性酶，存在于细胞外基质 (ECM) 和细胞核中，对于肺部的器官形态发生、组织修复和抗肿瘤发生至关重要。本实验室前期研究表明，镉（Cd）是一种有毒、致癌的重金属，可诱导大鼠肺成纤维细胞LO基因沉默。拟议的研究扩展了先前的发现，旨在研究细胞和动物模型中镉导致LO基因沉默的分子机制，为开发针对与镉相关的肺部疾病（例如肺气肿和癌症）的保护和治疗策略提供基础。