Proteome-scale programmable protein perturbation and induced proximity

Programmable perturbation technologies such as CRISPR-Cas have transformed how we study biological systems by offering efficient and scalable methods to manipulate DNA and RNA, thereby linking genotype to phenotype. However, corresponding technologies for rationally manipulating all proteins have lagged behind, restricting our ability to connect proteotypes to phenotypes. Developing a programmable protein perturbation technology would be a breakthrough in the biomedical sciences, as it would facilitate the interrogation of biology and creation of medicines in fundamentally new ways. The overarching goal of this proposal is to develop and apply such a high-throughput programmable protein perturbation platform by combining tools and concepts at the intersection of gene editing, proximity-induced pharmacology, and pooled screening. Our preliminary work has already established a foundation for this, including successful demonstration of high-throughput prime editing–mediated epitope tagging and targeted protein degradation (TPD) as well as induced proximity between any two proteins of interest. In this proposal, we will build upon these advances by: 1) establishing a proteome-scale TPD platform and applying it to uncover the time-resolved human protein essentialome, 2) interrogating the interactions between hundreds of therapeutic target proteins and thousands of protein degradation effectors in order to identify new strategies for induced-proximity therapeutics, and 3) characterizing a therapeutic landscape of cancer cell reprogramming strategies in which the function of cancer-associated proteins are overwritten by an endogenous effector protein. This proposal introduces new technologies for high-throughput programmable protein perturbation and induced proximity, offering insights into essential proteins, strategies for cancer treatment based on induced-proximity pharmacology, and a transformative methodology enabling broad applications in biology and medicine.