IntBIO: Chromosomal behavior underlying enhancer network function revealed by genomic perturbation, live fluorescent imaging, and theoretical modeling

This project seeks to uncover the fundamental physical rules that govern how genes are switched on and off within our cells. Inside the cell's nucleus, our meter-long DNA is intricately folded into 3-dimensional compartments, and this 3D organization is critical for controlling protein production. Special DNA segments called “enhancers” act as remote-control switches, physically looping through space to form contacts that activate specific genes, a process essential for normal development and cellular function. However, when this process goes awry, it can lead to diseases like cancer. This research will focus on MYC, a critical gene that, when over-activated, drives the growth of many cancers. By combining advanced genetic tools (CRISPR) with high-resolution live-cell microscopy, the research team will directly observe the movement of genomic DNA in real time inside living cells, monitoring the dynamic interactions between enhancers and the MYC gene. To decipher the complex rules of this dance, the team will use a combination of physics-based computer simulations and data-driven models using artificial intelligence. Understanding this process is a crucial step toward new strategies for treating genetic diseases. The project also advances the national interest by fostering scientific education through its LABScl program, which develops and delivers hands-on science activities for high school students in hospital schools and other non-traditional settings. This interdisciplinary research program will establish the causal relationship between chromosome dynamics and gene expression by integrating genomic perturbation, live-cell imaging, and theoretical modeling. The project focuses on the MYC oncogene locus, a prototypical system regulated by a network of enhancers. The research is structured in two aims. Specific Aim 1 will characterize the real-time, 3D dynamics of enhancer-promoter and enhancer-enhancer interactions within the MYC locus. The Qi lab will use Oligo-LiveFISH, a novel CRISPR-based imaging technology, to track multiple genomic loci simultaneously in living cells. These dynamics will be observed under normal conditions and following targeted perturbations of individual enhancers using CRISPR interference (CRISPRi). Concurrently, the Spakowitz lab will develop multi-scale polymer physics models and data-driven models using machine learning and artificial intelligence approaches, incorporating key biological factors, such as BRD4-induced chromatin condensation, to interpret the experimental trajectories. In Specific Aim 2, the team will determine how these measured dynamic interactions translate to gene expression outcomes. The experimental approach will be extended to visualize enhancer-promoter dynamics and the real-time transcription of MYC mRNA. To connect the system's biophysical parameters to its function, the team will develop a multi-layer perceptron (MLP), an Artificial Intelligence model, to predict enhancer-promoter contacts and gene expression from epigenetic profiles. This integrated experimental-computational framework, which advances both Biotechnology and AI, will provide fundamental insights into gene regulation and will be broadly transferable to other genetic systems. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria. NSF Award ID: 2611082 | Program: 01002627DB NSF RESEARCH & RELATED ACTIVIT | Principal Investigator: Andrew Spakowitz | Institution: Stanford University, STANFORD, CA | Award Amount: $1,000,000 View on NSF Award Search: https://www.nsf.gov/awardsearch/show-award/?AWD_ID=2611082 View on Research.gov: https://www.research.gov/awardapi-service/v1/awards/2611082.html