DREAM Mediated Transcription Acts as a Biomarker for Pollutant Induced Calcium Signaling Disruption

摘要/摘要 多氯联苯、多溴联苯醚和三氯生等化学品 改变通过L型电压敏感性Ca 2+通道（CaV 1）和/或兰尼碱作用的细胞Ca 2+信号传导 受体（RyR），其调节来自外部来源的Ca 2+的进入或来自内部来源的Ca 2+的释放。 商店，分别。这些通道本身就是大量细胞信号功能的核心 这些目标的过程和化学破坏与改变的树突状生长，改变的 神经肌肉健康，以及暴露的模式生物的学习和记忆改变。有毒物质清单，以及 它们的受体靶点，导致这种Ca 2+信号传导中断（CSD;用于中断和破坏）， 目前未知。然而，基于与已知CSD化合物的结构相似性，我们假设 目前正在使用或环境中的大量化学品能够通过CSD处理， 高通量工具能够筛选单个化学品和复杂的环境混合物。最近的工作已经 鉴定了下游调节元件拮抗调节剂（DREAM），其是唯一的Ca 2+传感器 迄今为止，已鉴定出特异性结合DNA并以Ca 2+依赖性方式调节转录。因此 DREAM蛋白在细胞内Ca 2+变化和基因转录之间起着直接联系的作用 我们的初步工作表明，选择CSD化合物可以改变DREAM介导的转录， 代表建立CSD筛选测定的独特机会的模型细胞系。 在这里，我们采取多层次的方法来扩展我们对梦想的理解，根据污染物的反应， 通过筛选已知和预测的CSD化学物质，以确定它们驱动荧光素酶中DREAM转录的能力， 转染的细胞系，并评估DREAM介导的转录或CSD中改变的表型的程度 暴露的野生型和DREAM敲除斑马鱼。这项拟议中的工作将建立基因转录作为一种有毒物质， 由于CSD的结果，并导致重要的细胞和动物工具的开发和验证， 成本和时间有效地评估大量化合物的CSD活性。这项工作具有广泛的意义 对暴露生物体的健康的影响，因为众所周知，梦有助于学习和 下丘脑、垂体和甲状腺中的记忆、疼痛接收和内分泌信号， 与亨廷顿病有关， 肌萎缩侧索硬化 和II型糖尿病理解 CSD在这些生理和病理生理状态中作用将有助于与污染物相关的风险 考核

Summary/Abstract Chemicals such as polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs) and triclosan alter cellular Ca2+ signaling acting through L-type voltage-sensitive Ca2+ channels (CaV1) and/or ryanodine receptors (RyR), which regulate the entry of Ca2+ from external sources or the release of Ca2+ from internal stores, respectively. These channels alone are central to the function of a vast number of cellular signaling processes and chemical disruption of these targets is correlated with altered dendritic growth, altered neuromuscular health, and altered learning and memory in exposed model organisms. The list of toxicants, and their receptor targets, that lead to such Ca2+ signaling disruption (CSD; used for disruption and disrupting) is currently unknown. However, based on structural similarities to known CSD compounds, we hypothesize that a large number of chemicals currently in use or in the environment are capable of CSD representing a need for high-throughput tools able to screen individual chemicals and complex environmental mixtures. Recent work has identified the downstream regulatory element antagonistic modulator (DREAM) which is the only Ca2+ sensor identified to date that binds specifically to DNA and regulates transcription in a Ca2+-dependent manner. Thus the DREAM protein acts as a direct connection between changes in intracellular Ca2+- and gene transcription Our preliminary work demonstrates that select CSD compounds can alter DREAM mediated transcription in model cell lines representing a unique opportunity to establish a CSD screening assay. Here, we take a multi-tiered approach to extend our understanding of DREAM, in-light-of pollutant responses, by screening known and predicted CSD chemicals for their ability to drive DREAM transcription in luciferase transfected cell lines and evaluating the extent of DREAM mediated transcription or altered phenotypes in CSD exposed wildtype and DREAM knockout zebrafish. The proposed work will establish gene transcription as a toxic outcome due to CSD and lead to the development and validation of important cellular and animal tools able to cost and time effectively evaluate large numbers of compounds for CSD activity. This work has vast implications on the health of exposed organisms as DREAM is known to contribute to such processes as learning and memory, pain reception and endocrine signaling in the hypothalamus, pituitary and the thyroid gland and has been associated with Huntington’s disease, a myotrophic lateral sclerosis and type II diabetes. Understanding the role of CSD in these physiological and pathophysiological states would aid in pollutant associated risk assessment.