FANCONI ANEMIA GENE PATHWAY IN RADIATION RESPONSES

该项目的目标是了解细胞用于减少内源性过程或电离辐射（IR）引起的活性氧（ROS）相关的遗传损伤和遗传不稳定性的分子调控过程。这一目标是通过FANCG/XRCC9的研究来解决的。癌症易感性疾病范可尼贫血（FA）的G组。由于FancG蛋白在仓鼠细胞中具有IR抗性，因此人类同源物有望参与人类细胞的IR反应。从历史上看，FA基因与辐射反应之间的联系尚不清楚，一些研究表明FA的主要缺陷在于去除DNA链间交联。需要验证的一般假设是，FANCG蛋白作为多蛋白复合物的一员，保护哺乳动物细胞免受内源性和红外产生的氧化损伤，并通过协调包括ROS水平调节、细胞凋亡和细胞周期进程在内的稳态过程来维持基因组完整性。拟议的研究将提供FANCG蛋白对与正常细胞增殖和对IR暴露反应相关的生化和细胞终点的贡献的高度定量表征。将在仓鼠CHO细胞和人淋巴母细胞中获得突变细胞和fancg补充细胞的等基因对。这些染色体对将分析染色体畸变、细胞存活、hprt基因突变、细胞凋亡、ROS和细胞周期参数是否有IR暴露。fancg补充FA-G淋巴母细胞将用于检查细胞周期中的基因和蛋白质调控以及有和没有IR损伤的蛋白质的亚细胞定位。从初步研究中获得的三种候选FANCG相互作用蛋白将被评估是否可能参与FA通路。最后，在美国人群中已经确定的FANCG高频人类等位基因变异将被评估其功能障碍程度。这些研究的结果将导致FA蛋白“途径”性质的更具体模型及其对与ir介导的氧化损伤相关的多种生物效应的定量贡献。

The project's objective is to understand the molecular regulatory processes cells use to minimize genetic damage and genetic instability associated with reactive oxygen species (ROS) arising from endogenous processes or ionizing radiation (IR). This goal is addressed through studies of FANCG/XRCC9, the gene that is defective in. group G of the cancer-prone disorder Fanconi anemia (FA). Because FancG protein confers IR resistance in hamster cells, the human homolog is expected to participate in IR responses in human cells. Historically, a link between the FA genes and radiation responses has been unclear, with some studies suggesting that the primary defect in FA lies in removing DNA interstrand crosslinks. The general hypothesis to be tested is that the FANCG protein, as a member of a multiprotein complex, protects mammalian cells against endogenous and IR-generated oxidative damage and maintains genomic integrity by coordinating homeostasis processes that include regulation of ROS levels, apoptosis, and cell cycle progression. The proposed studies will provide a highly quantitative characterization of FANCG protein's contribution to biochemical and cellular endpoints associated with both normal cell proliferation and responses to IR exposure. Isogenic pairs of mutant and FANCG-complemented cells will be derived in both hamster CHO cells and human lymphoblasts. These pairs will be analyzed with respect to chromosomal aberrations, cell survival, hprt gene mutations, apoptosis, ROS, and cell cycle parameters with and without IR exposure. The FANCG-complemented FA-G lymphoblasts will be used to examine gene and protein regulation during the cell cycle as well as the subcellular localization of the protein with and without IR damage. Three proteins that are candidate interactors with FANCG from preliminary studies will be evaluated for possible involvement in the FA pathway. Finally, already identified high-frequency human allelic variants of FANCG in the US population will be evaluated for degree of dysfunction. The results of these studies will lead to more specific models of the nature of the FA protein "pathway" and its quantitative contributions to multiple biological effects associated with IR-mediated oxidative damage.