Endogenous Gas Molecules As Transcription Factors

描述（由申请人提供）：气体分子在哺乳动物生存中的重要性可以通过氧气维持生命的必要性来证明。2000年，诺贝尔医学奖被授予发现和鉴定一氧化氮（NO）作为内皮来源的松弛因子。这一发现彻底改变了我们对气体分子的看法，并摒弃了许多实验室研究NO调节细胞功能的分子机制的活动，特别是在心血管领域。最近在1998年，发现了第二个功能气体分子；一氧化碳，被吹捧为一种应该避免的毒素，被证明是一种生物活性气体分子，在体外和体内低浓度下具有有效的细胞保护特性。这一发现源于对血红素加氧酶-1的研究，这是一种诱导应激反应基因，在所有细胞中分解血红素，内源性产生一氧化碳。CO和NO已经得到了很好的研究，并将继续在许多模型系统中从机制的角度进行评估。关于这些气体与信号级联和下游基因调控相关的作用，已经收集了大量的信息。与NO不同，CO是非反应性的，它作用于许多酶的血红素部分中含有的二价阳离子，如铁，以调节其功能。CO是否可能与其他金属阳离子结合，但尚未研究。在阐明这些气体作为生物介质之前，气体一直被认为是满足细胞代谢需求所必需的，或者是酶促过程的简单废物。我们相信气体在整个细胞功能和行为中发挥着更大的作用，并提出了一种创新的假设，即我们将使用CO作为我们研究的原型气体，在DNA动力学水平上作为气体转录调节剂在所有细胞中作为稳态传感器运行。我们相信这符合EUREKA机制的目的，因为它是一种新颖的创新和非常规的假设，如果被证明是有效的，将重塑当前的DNA调控理论，以及细胞功能和行为的许多方面，包括基因表达，但可能更重要的是DNA损伤和修复，以及DNA合成和增殖。我们的中心假设是气态CO通过金属离子与DNA上存在的聚合酶和拓扑异构酶的复合物直接与DNA相互作用。在分配给这项工作的时间里，我们将评估CO与DNA的相互作用，以及它如何影响细胞中的转录、损伤识别和增殖。我们将以以下目标来验证我们的假设：具体目标1：测试CO调节DNA动力学的能力。具体目标2：评估CO与DNA和/或聚合酶结合的后果，通过促进DNA解绕和促进聚合酶活性来调节转录。特异性目的3：探讨CO在DNA合成和细胞增殖调控中的作用。公共卫生相关性：了解细胞如何控制自己的命运并对环境作出反应，对于科学发现如何干预和纠正不适当的反应或细胞功能的变化至关重要，这些变化是疾病病理起源的基础。我们提出气体一氧化碳（CO）是由所有细胞内源性产生的，是一种通过影响DNA如何调节基因表达直接影响细胞行为的分子。低、无毒浓度的一氧化碳在动物疾病模型中具有有效的保护和修复作用。这个提议将集中在一氧化碳和一个创新的假设，即气体分子可以在DNA水平上指示细胞反应。

DESCRIPTION (provided by applicant): The importance of gas molecules in mammalian survival is exemplified by the requisite need for oxygen to sustain life. In 2000 the Noble Prize in Medicine was awarded for the discovery and characterization of nitric oxide (NO) as endothelial derived relaxation factor. This discovery revolutionized how we view gas molecules and spurned the activity of numerous laboratories to study the molecular mechanisms by which NO regulated and modulated cellular function, particularly in the cardiovascular arena. More recently in 1998, a second functional gas molecule was revealed; carbon monoxide, touted as a toxin to avoid, was shown to be a biologically active gas molecule with potent cytoprotective properties in vitro and in vivo at low concentrations. This discovery stemmed from work on the enzyme heme oxygenase-1, which is an inducible stress response gene that generates CO endogenously as it catabolizes heme in all cells. CO and NO have been well studied and continue to be evaluated from a mechanistic standpoint in numerous model systems. A great deal of information has been gleaned regarding the action of these gases related to signaling cascades and downstream gene regulation. CO unlike NO is non-reactive and acts on divalent cations such as iron contained in heme moieties of numerous enzymes to modulate their function. Whether CO binds to other metal cations is likely but has not yet been studied. Until the elucidation of these gases as biological mediators, gases have carried the dogma of simply being necessary to either fulfill metabolic requirements of the cell or as simple waste products of enzymatic processes. We believe there is a greater role for gases in overall cellular function and behavior and offer the innovative hypothesis that CO, which we will use as the prototype gas for our studies, functions as a gaseous transcriptional regulator operating as a homeostatic sensor within all cells at the level of DNA dynamics. We believe this fits the purpose of the EUREKA mechanism because it is a novel innovative and unconventional hypothesis which if proven to be valid will reshape current theory of DNA regulation, but also many aspects of cellular function and behavior including gene expression, but perhaps more importantly DNA damage and repair, as well as DNA synthesis and proliferation. Our central hypothesis is that gaseous CO interacts directly with DNA via metal ions in complex with polymerases and topoisomersases present on DNA. In the time allotted for this work we will evaluate the interaction of CO with DNA and how this influences transcription, recognition of damage and proliferation in the cell. We will test our hypothesis with the following aims: Specific Aim 1: To test the ability of CO to modulate DNA dynamics. Specific Aim 2: To evaluate the consequences of CO binding to DNA and/or polymerase to regulate transcription by fostering the unwinding of DNA and facilitating polymerase activity. Specific Aim 3: To evaluate the role of CO in DNA synthesis and the regulation of cellular proliferation. PUBLIC HEALTH RELEVANCE: Understanding how a cell controls its own destiny and responds to its environment is critical to scientific discoveries of how to interfere and correct an inappropriate response or change in the cell function that underlie the origins of disease pathology. We are proposing that the gas carbon monoxide (CO), which is generated endogenously by all cells, is a molecule that directly influences cellular behavior by influencing how DNA is regulated for gene expression. Low, non-toxic concentrations of CO impart potent protection and repair in animal models of disease. This proposal will focus on carbon monoxide and the innovative hypothesis that a gas molecule can dictate a cellular response at the level of the DNA.