Super-resolution microscopy for dynamic analysis of focal enhancer amplifications in cancer

项目摘要 癌症通常被认为是一种遗传性疾病。例如，特定的蛋白质编码突变可以 激活致癌基因。然而，致癌基因也可以通过改变 非编码DNA具体来说，增强子是可以激活基因表达的非编码DNA元件 跨越巨大的基因组距离。在许多癌症中，基因改变导致增强子过度激活， 致癌基因这也被称为增强器劫持。 在这里，我们集中在上皮癌，其中激活MYC癌基因的增强子得到复制 并聚焦放大。这与较高的MYC表达和增加的致瘤性相关。但 分子机制还不清楚。实现机械的理解是重要的，因为它可能 确定分子的“阿喀琉斯之踵”，可以针对治疗。 理解增强子的一般作用机制和增强子劫持， 癌症通常需要以高时空精度测量增强子-基因通讯的能力 在活细胞中。拟议的项目将建立和验证这样一个基于超分辨率的平台 显微镜，这使得有可能跟踪个人增强子和基因的精度为10纳米 活细胞中的基因活动以及真实的时间。将投入大量精力来验证超级 分辨率显微镜方法通过广泛的控制实验。还将作出重大努力， 向更大的社区传播技术创新和知识。 总之，我们建议建立和验证一个创新的和综合的实验和 计算超分辨率显微镜平台，用于研究焦点增强器放大。虽然我们最初的 重点是上皮癌中MYC增强子的扩增，所提出的平台可能会发现 广泛使用超过MYC。

Project Summary Cancer is often thought of as a genetic disease. For example, specific protein-coding mutations can activate cancer-causing oncogenes. However, cancer-causing oncogene can also be activated by alterations to non-coding DNA. Specifically, enhancers are non-coding DNA elements that can activate gene expression across vast genomic distances. In numerous cancers, genetic alterations cause enhancers to over-activate cancer-causing oncogenes. This is also referred to as enhancer hijacking. Here we focus on epithelial cancers, where enhancers that activate the MYC oncogene get duplicated and focally amplified. This correlates with higher MYC expression and increased tumorigenicity. But the molecular mechanisms are not well understood. Achieving a mechanistic understanding is important since it may identify molecular ‘Achilles Heels’ that can be targeted therapeutically. Achieving an understanding of the mechanism of enhancer action in general and enhancer hijacking in cancer in general requires an ability to measure at high spatiotemporal precision enhancer-gene communication in living cells. The proposed project will establish and validate such a platform based on super-resolution microscopy, which make it possible to track individual enhancers and genes with a precision of ten nanometers in living cells as well as gene activity in real time. Significant efforts will be devoted to validating the super- resolution microscopy approach through extensive control experiments. Significant efforts will also be devoted to disseminating the technological innovations and know-how to the greater community. In summary, we propose to establish and validate an innovative and integrative experimental and computational super-resolution microscopy platform for studying focal enhancer amplifications. While our initial focus is on amplifications of MYC enhancers in epithelial cancers, the proposed platform is likely to find widespread use beyond MYC.