Protein-ligand interface engineering for allele-specific regulation of histone demethylases and epigenome editing

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In eukaryotes, genes can be turned on or off by changing the way the genome is packaged in the nucleus. Normally, the genome is compacted by winding the DNA around barrel-like cores of proteins. Tight packaging turns genes off, but chemical modification of the protein cores can relax the DNA and allow genes to be turned on. The modifications are carried out by a large group of enzymes, but how and where they function remains unknown. This project will take an interdisciplinary approach, including organic chemistry, protein engineering, and cell biology--to uncover the specificity of these modifiers and to design enzymes capable of turning on specific genes at will. The research setting will provide a unique training ground for graduate and undergraduate students. Furthermore, a chemical biology laboratory course will engage undergraduate students in inquiry-based exercises where students will learn to design and carry out real-world experiments aimed at developing their critical thinking and independent learning skills. Early implementation of such a research-based course is expected to bridge a gap existing in the current educational curriculum by motivating STEM students in science education and research with the long-term benefit of generating a skilled workforce.Changes in gene expression in eukaryotic organisms like humans can be achieved by reversible chemical modifications on the histone protein components of chromatin. The focus of this project is to study a particular type of modification, called lysine methylation, which is removed by a class of enzymes known as lysine demethylases. How specific demethylases contribute to gene expression has remained largely unexplored, due in part to the lack of tools capable of rapidly interrogating a given demethylase in intact cells under carefully controlled conditions. Using one particular lysine demethylase as paradigm, this research will focus on developing a novel chemical-genetic platform that combines pharmacological and genetic engineering to perturb specific isoforms with rationally designed small molecules and precise temporal control. Furthermore, the engineered demethylation apparatus will be combined with the spatial selectivity of programmable CRISPR-Cas9 to develop a new type of conditional epigenome editing tool for regulating gene transcription in space and time. The approach will be applied to reprogram expression of genes that underlie faithful cell division, cellular differentiation, lineage commitment, and ultimately, organismal development. These unique tools will be made broadly available to researchers interested in addressing how reversible histone methylation regulates eukaryotic biology.This project is funded jointly by the Genetic Mechanisms Cluster, Division of Molecular and Cellular Biosciences in the Directorate of Biological Sciences and the Chemistry of Life Processes Program, Division of Chemistry in the Directorate of Mathematical and Physical Sciences.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.