CAREER: A Synthetic Biology Platform to Map and Engineer the Diverse Epigenetic Space

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This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2).This project will develop new technological platforms and educational innovations that will unlock our understanding of and ability to control the expression of genes in higher order organisms from yeast and plants to humans, with broad applications in biotechnology. Chromatin is the layer of hundreds of proteins bound on top of the genome. It confers regulatory handles over diverse cellular and organismal processes including aging and cancer, and it could be leveraged in biotechnological applications including bioremediation, environmental sensing, and bio-based production of commodity chemicals. Chromatin also provides a strong topic for pedagogical innovation and enrichment. This project will develop hands-on experiments for middle and high school students that connect molecular changes in chromatin to effects observable by eye, in this case the binding of yeast cells to the surface of an agar plate. Furthermore, the numerical and biochemical diversity of chromatin advocates for new pedagogical approaches at the collegiate and PhD levels. Specifically, this project will develop new coursework modules that train the next generation of scientists in how to handle and analyze large data sets. American Rescue Plan funding of this project will support the investigator at a critical stage in his career.Chromatin is incredibly diverse at the molecular level. There are more than 80 different amino acid residues on five histone proteins known to undergo over 20 distinct post-translational modifications. Hundreds of proteins, often containing domains referred to as writers and readers, both regulate and interact with this complex biochemical palette. Through the development of high content technologies paired with biochemical and genetic approaches it has become clear that much of this diversity is functional. However, unlike many methods that can characterize chromatin at high throughput, functional assays remain substantially more limited in scale. In particular, characterizing and engineering the enzymatic activities and specificities of epigenetic writers still rely largely on low throughput biochemical assays that require recombinant protein production and purification that are challenging to scale even with robotic liquid handling. This proposal seeks to build upon recent synthetic biology platforms the investigator and his team have developed using yeast surface display and epigenome editors to unlock access to the diversity of chromatin. With a focus on histone acetyltransferases (HAT), they will establish proof-of-principle demonstrations that these systems can 1) map the residue specificities of all known human HATs; 2) connect sequence to function by accessing the very large enzyme and substrate sequence space by directed evolution approaches; and 3) probe the impact of molecular context on function. While the investigations focus on HATs, the investigator and his team seek to motivate the future expansion and application of this experimental platform to the diverse biochemistry of chromatin.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.