Epithelial stress sensors: novel roles for cytochrome P450s and organellar calcium in integrated stress and immune responses.

所有细胞都会因化学反应产生活性氧(ROS)，这也发生在细胞内的细胞器中。例如，线粒体产生的能量和过氧化物体中的化学反应产生ROS。细胞和组织使用不同的酶和过程来处理ROS的产生，这些酶和过程‘中和’ROS。当ROS的产生与相关细胞或组织处理带电离子的能力之间存在不平衡时，就会产生“氧化应激”。细胞对氧化应激的反应是由信号转导机制调节的，信号转导机制利用包括钙离子(Ca~(2+))在内的小信号分子。氧化应激影响细胞蛋白质、脂质和DNA，在人类中与神经退行性疾病、包括中风和心脏病发作在内的心血管疾病以及衰老过程有关。然而，ROS的产生也与免疫密切相关，因为ROS可用于防御感染。人们越来越认识到，氧化应激机制和免疫功能是联系在一起的，这是生物学机制的基本原理以及人类健康和老龄化的重要研究领域。无脊椎动物模式生物，如黑腹果蝇，是研究基因功能在生物应激反应中的影响的中心。此外，对果蝇的研究也提供了操纵压力相关基因和生理输出之间最明确的联系。应激反应基因的组织或细胞特异性背景在组织对应激反应中的重要性使果蝇特别有用，因为在果蝇中对基因的细胞和组织特异性靶向是众所周知的。果蝇上皮马氏(肾)管相当于脊椎动物的肾脏和肝脏，在解毒、液体和离子运输中起着关键作用。小管充满了线粒体和过氧化物体，并产生ROS作为其正常功能的一部分。我们已经证明，小管具有特定的适应能力，以对抗线粒体的高活性和ROS的产生，包括富含“抗氧化”基因，这些基因在人类中也是保守的。我们还表明，过量的ROS对小管线粒体钙信号的去调节会导致小管细胞死亡(细胞凋亡)和整个苍蝇的死亡。因此，这种上皮组织在整个生物体的压力感应中起着核心作用。小管也是一个独立的免疫感应组织，它使用“天生”的免疫机制来感知细菌或真菌的挑战。上皮细胞中的这种天然免疫机制在脊椎动物中也是保守的，并且构成了一个日益重要的研究领域，因为现在已经知道，包括克罗恩氏病和哮喘在内的疾病是由于这些液体运输的上皮组织(分别是肠和肺)的天然免疫机制的放松调节所致。我们的最新数据来自对氧化应激和免疫挑战的生存研究，对线粒体钙信号的研究，以及对苍蝇应激反应的~13500个基因的分析表明，关键的新的小管特异基因参与了苍蝇的应激/免疫反应。这些基因编码“解毒”蛋白--细胞色素P450，通常与线粒体或过氧化物体相关，在这里，我们显示了与应激反应有关。我们将在过氧化酶体和线粒体中钙信号的背景下研究这些基因，并使用新基因的功能分析、遗传学/转基因技术来评估这些基因/蛋白质对整个生物体生存的影响，钙报告程序针对完整组织中的线粒体和过氧酶体。这将使我们能够发现调节上皮细胞应激反应的新方法，这可能会对人类生物学产生影响。

All cells produce reactive oxygen species (ROS) due to chemical reactions, which also occur in the organelles contained within cells. For example, the production of energy from the mitochondria and chemical reactions in the peroxisomes generate ROS. Cells and tissues deal with the production of ROS using different enzymes and processes, which 'neutralise' ROS. When there is an imbalance between the production of ROS and the ability of the cells or tissues in question to deal with the charged ions, 'oxidative stress' results. Cellular responses to oxidative stress are regulated by signal transduction mechanisms which utilise small signalling molecules including calcium ions (Ca2+). Oxidative stress affects cellular proteins, lipids and DNA, and in humans is associated with neurodegenrative disease, cardiovascular disease including stroke and heart attack, and with the ageing process. However, ROS production can also closely linked to immunity, as ROS can be used in defence against infection. It is increasingly known that mechanisms of oxidative stress and immune function are linked, and these are important areas of research for the basic fundamentals of biological mechanisms, and for human health and ageing. Invertebrate model organisms eg., Drosophila melanogaster, are central for investigating the impact of gene function in stress responses in the organism. Furthermore, studies in Drosophila have also have provided the clearest links between manipulation of stress-associated genes and physiological output. The importance of the tissue- or cell-specific context of stress-response genes in oragnismal responses to stress, makes Drosophila particularly useful, as cell- and tissue-specific targeting of genes in Drosophila is well-established. The Drosophila epithelial Malpighian (renal) tubule is equivalent to vertebrate kidney and liver and plays critical roles in detoxification, fluid and ion transport. Tubules are packed with mitochondria and peroxisomes, and produce ROS as part of their normal function. We have shown that tubules have specific adaptations to counter the high mitochondrial activity and production of ROS, including being enriched for 'anti-oxidant' genes which are also conserved in humans. We have also shown that de-regulation of tubule mitochondrial calcium signalling by excess ROS leads to tubule cell death (apoptosis) and death of the whole fly. Thus, this epithelial tissue has a central role in stress-sensing for the whole organism. The tubule is also a stand-alone immune-sensing tissue which uses 'innate' immune mechanisms to sense bacterial or fungal challenge. Such innate immune mechanisms in epithelia are also conserved in vertebrates, and constitute an increasingly important area of study, as it is now known that diseases including Crohns' Disease and asthma are due to deregulation of innate immune mechanisms in these fluid-transporting epithelial tissues (intestine, and lungs, respectively). Our recent data from survival studies in response to oxidative stress and immune challenge, work on calcium signals in mitochondria and analysis of the ~13500 genes of the fly in response to stress shows that key novel tubule-specific genes involved with the stress/immune responses the fly. These genes encode 'detoxification' proteins-the cytcochrome P450s, normally associated with mitochondria or peroxisomes and here, we have shown involvement with the stress response. We will investigate these genes in the context of calcium signals in the peroxisome and mitochondria, and assess the impact of each of these genes/proteins on survival of the whole organism using functional assays for novel genes, genetics/transgenics with calcium reporters targeted to mitochondria and peroxisomes in the intact tissue. This will allow us to uncover new ways of regulating stress responses in epithelial cells, which may impact on human biology.