Novel markers of exposure and pathways of response to Chromium

摘要 铬(VI)是一种引起重大公共卫生问题的人类致癌物质，在许多 职业设置。铬(VI)诱发突变，改变基因拷贝数，并暴露于 与暴露人群和动物模型中的人类癌症有关。我们新奇的初步数据 证明铬(VI)暴露可引起核糖体DNA(RDNA)拷贝数的放大和 核仁(对核糖体RNA(RRNA)转录位置的粗略理解的核细胞器， 以及无数细胞功能的集成)。核仁功能的一个关键因素是rDNA拷贝数 (RDNA CN)。RDNA CN调节(I)整个基因组的表观遗传状态，(Ii)DNA损伤反应，(Iii)细胞 周期进程、(Iv)染色体分离和(V)全球遗传稳定性。此外，中断 RDNA阵列、核糖体生物发生和核仁是癌变的中心。我们的中央医疗中心 假设铬诱导的rDNA CN的改变是铬致癌的原因。我们的中央 基本假设是rDNA阵列不是固定的，而是核的遗传动态成分 基因组的拷贝数受铬暴露的影响。我们的建议是检查rDNA的变化 铬(VI)暴露揭示了一种新的铬毒性途径，具有医学和基础相关性。关键要素是 假设驱动的铬诱导rDNA扩增(Cr-I-rDNA-a)毒理学研究 铬(VI)暴露下rDNA和核仁的功能基因组分析和广泛的遗传分析 利用一种强大的模式生物对铬-i-rDNA-a进行了研究。我们的首要目标是研究铬的毒理学-- 人肺上皮细胞模型的诱导rDNA扩增(Cr-I-rDNA-a)。我们将确定剂量- 铬-i-rDNA-a的反应，在铬-i-rDNA-a中的MAP扩增边界，检查时间分布和 从铬(VI)暴露中恢复，并检查铬诱导的CN变化是否对铬- 诱导致癌。我们的第二个目标是研究铬诱导核仁应激的功能基因组学 和铬在人肺上皮细胞模型中诱导转化。检测全基因组对铬的反应 接触铬对了解铬如何诱导rDNA扩增、核应激和致癌是至关重要的。我们的 第三个目标是解决铬-rDNA相互作用的遗传决定因素。我们将检测铬-i-rDNA-a 特定细胞，量化拷贝数变化的程度，分离受影响的组织，并使用高通量 描述这些变化的技术。我们的努力将阐明铬-rDNA的相互作用，研究 与美国国立卫生研究院的人类健康使命直接相关。RDNA CN的多种效应表明 用铬干扰这个中央调节器将对细胞功能产生深远的影响。我们期待着 癌症等具有强烈环境因素的复杂人类疾病的决定因素将 最终被追溯到环境引发的基因组rDNA片段的变异。

Abstract Cr(VI) is a human carcinogen of significant public health concern, and a substantial exposure in a number of occupational settings. Cr(VI) induces mutations, changes in gene copy number, and exposure has been associated with humans cancers in exposed populations and animal models. Our novel preliminary data demonstrate that Cr(VI) exposure causes amplification in ribosomal DNA (rDNA) copy number and changes in the nucleolus (the crudely understood nuclear organelle that is the site of ribosomal RNA (rRNA) transcription, and integration of myriad cellular functions). A crucial element of nucleolar function is rDNA copy number (rDNA CN). rDNA CN modulates (i) epigenetic states across the genome, (ii) DNA damage responses, (iii) cell cycle progression, (iv) chromosome segregation, and (v) global genetic stability. Furthermore, disruption of rDNA arrays, ribosome biogenesis, and the nucleolus are central to carcinogenesis. Our central medical hypothesis is that Cr-induced changes in rDNA CN are responsible for Cr-induced carcinogenesis. Our central basic hypothesis is that rDNA arrays are not fixed, but rather a genetically dynamic component of the nuclear genome with copy number that is modulated by Cr exposure. Our proposal examines rDNA changes upon Cr(VI) exposure to reveal a novel pathway of Cr toxicity with medical and basic relevance. Key elements are a careful investigation of the toxicology of Cr-induced-rDNA-amplification (Cr-i-rDNA-a), hypotheses-driven functional genomic analysis the rDNA and the nucleolus upon Cr(VI) exposure, and extensive genetic analyses of Cr-i-rDNA-a using a powerful model organism. Our first aim will investigate the toxicology of Chromium- induced-rDNA-amplification (Cr-i-rDNA-a) in a human lung epithelial cell model. We will determine dose- responses of Cr-i-rDNA-a, map amplification boundaries in Cr-i-rDNA-a, examine temporal profiles and recovery from Cr(VI) exposure, and examine whether Chromium-induced CN changes are responsible for Cr- induced carcinogenesis. Our second aim investigates the functional genomics of Cr-induced nucleolar stress and Cr induced transformation in a human lung epithelial cell model. Examining genome-wide responses to Cr exposure is critical to understand how Cr induces rDNA amplification, nuclelar stress, and carcinogenesis. Our third aim addresses the genetic determinants of Chromium-rDNA interactions. We will examine Cr-i-rDNA-a in specific cells, quantify the extent of copy number change, isolate the affected tissues, and use high-throughput techniques to characterize the changes. Our efforts will shed light on Cr-rDNA interactions, with research that is directly relevant to the human health mission of the NIH. The manifold effects of rDNA CN indicate that perturbing this central regulator with Cr will have profound consequences to cellular function. We anticipate that determinants of complex human diseases with strong environmental components such as cancer will ultimately be traced to environmentally triggered variation in rDNA segments of the genome.