Exploring arsenic and its metabolites in a transgenic model

描述（由申请人提供）：全球公共卫生界面临的最严重问题之一是饮用水中摄入砷的长期后果，这一情况已影响到全世界约1亿人，尽管在孟加拉国和西孟加拉邦受到的影响尤其严重，因为广泛提供的水井抽取含砷地下水。虽然砷是一种众所周知的氧化磷酸化急性毒物，当大量摄入时，慢性，低水平暴露会导致各种病理，其中包括多种形式的癌症。从机理上讲，很难确定砷诱发癌症的具体原因，尽管DNA和/或染色体畸变通常伴随着它的出现，而且关于砷是否可以作为一种完全的致癌物一直存在很多争论。此外，由于相对缺乏合适的动物模型，这些领域的进展受到阻碍。最近，人们认识到砷的代谢甲基化实际上可能导致一种更强的致癌物，而不是其假定的解毒作用，这促使人们对甲基化的机制、形成的物种以及参与砷摄取、代谢和排泄的基因多态性的可能性产生了兴趣，这些基因多态性可能对暴露个体的易感性产生了深远的影响。为了用一个定义良好的、遗传上可适应的体内系统来解决这些问题，我们建议创建一个果蝇转基因模型，在这个模型中，砷甲基化的控制和精确分析（由人群中自然发生的人类基因变异催化）可以与各种体内分析相结合，研究DNA代谢的特定特征（氧化损伤、链断裂、重组）、细胞反应（染色体畸变、细胞周期畸变）、以及在移植试验中通过肿瘤形成的致癌潜力。由于果蝇系统具有巨大的通用性，可以很容易地实现几乎任何内源性基因的可控过表达和过表达，因此可以预期，砷及其甲基化代谢物交叉的关键分子途径可以在上述一种或多种检测中被识别为表型变异的结果，当在这种改变的遗传背景下进行测试时。因此，我们提出一个模型高等真核生物，一个一直被证明是一个无与伦比的资源，在揭示许多人类病理条件的关键分子特征，可以利用来阐明一个至关重要的毒理学问题。公共卫生相关性：世界许多地区的人口通过饮用水长期摄入砷已被证明是现代最大的全球公共卫生灾难之一，因为随之而来的各种有害健康后果，包括多种形式的器官癌症。尽管从培养细胞的研究中获得了很多兴趣，但由于相对缺乏复制人类病理的良好动物模型，对这种有毒金属影响生物途径的机制（特别是其致癌能力）的调查一直受到阻碍。我们的目标是通过将参与砷代谢的关键人类基因引入果蝇来开发砷细胞毒性和肿瘤发生的模型，在果蝇中，结合这种生物体无与伦比的遗传可操控性，结合各种检测，预计将为砷如何诱发癌症以及为什么个体对其影响表现出广泛不同的易感性等问题提供新的线索。

DESCRIPTION (provided by applicant): Among the most serious issues confronting the global public health community is the long term consequence of arsenic ingestion in drinking water, a situation that has affected an estimated 100 million people worldwide, albeit disproportionately so in Bangladesh and West Bengal owing to the widespread provision of wells drawing arsenic-laced groundwater. Though arsenic is a well-known acute poison of oxidative phosphorylation when ingested in large quantities, chronic, low-level exposure leads to a variety of pathologies, among which are numerous forms of cancer. Mechanistically it has proven difficult to assign the specific cause of arsenic-induced cancer, though DNA and/or chromosomal aberrations typically accompany its appearance, and there has been much debate regarding whether or not arsenic can act as a complete carcinogen. Moreover, progress in these areas has been hampered by a relative lack of suitable animal models. Recently, the realization that metabolic methylation of arsenic may actually lead to a more potent carcinogen rather than to its presumed detoxification has prompted interest in the mechanism of methylation, the species formed, and the possibility that polymorphisms in the gene(s) involved in uptake, metabolism and excretion of arsenic may have profound effects on susceptibility of exposed individuals. To address some of these issues with a well-defined, genetically amenable, in vivo system we propose to create a Drosophila transgenic model, in which control, and precise analysis, of arsenic methylation (catalyzed by human gene variants that occur naturally in the population) can be married with a variety of in vivo assays investigating specific features of DNA metabolism (oxidative damage, strand breakage, recombination), cellular response (chromosomal aberrations, cell cycle aberrations), and carcinogenic potential via tumor formation in a transplantation assay. With the enormous versatility available in the Drosophila system, allowing controllable over- and underexpression of virtually any endogenous gene to be easily achieved, it is anticipated that critical molecular pathways intersected by arsenic and its methylated metabolites can be identified as a result of phenotypic variation occurring in one or more of the above assays when tested in such altered genetic backgrounds. Thus, we propose that a model higher eukaryotic organism, one that has consistently proved to be an unparalleled resource in uncovering critical molecular features of numerous human pathological conditions, can be harnessed to shed light on a vitally important toxicogenetic problem. PUBLIC HEALTH RELEVANCE: The long-term ingestion of arsenic via drinking water by human populations in many parts of the world has proven to be one of the largest global public health disasters of modern times owing to the variety of detrimental health consequences that ensue, including numerous forms of organ cancer. Though much of interest has been learned from studies in cultured cells, investigation of the mechanisms by which this toxic metal affects biological pathways (particularly in its ability to cause cancer) has been hampered by a relative lack of good animal models that duplicate the human pathologies. We aim to develop a model for arsenic cellular toxicity and tumorigenesis by introducing critical human genes involved in arsenic metabolism into the fruit fly, Drosophila, where a combination of assays, allied to the unparalleled genetic manipulability of this organism, are anticipated to shed new light on questions related to how arsenic induces cancer and why individuals show widely variable susceptibility to its effects.