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.

Project Narrative Cancer-causing oncogenes are often regulated by distal non-coding DNA elements called ‘enhancers’. In many cancers, the enhancer element gets duplicated and amplified which correlates with higher oncogene expression. However, the mechanisms through which enhancer amplification causes oncogene overexpression are not well understood and we therefore propose to establish and validate super-resolution microscopy techniques to address this gap in our understanding.