Decipher in situ signaling and complex genetics with cellular recording and combinatorial perturbations

Abstract

The complex, dynamic, and responsive behavior of cells arises from integrated signaling pathways and regulatory networks. With advancement in our ability to engineer mammalian cells, we harness a novel set of molecular tools to develop synthetic biology-enabled applications that help facilitate our understanding of complex biological networks and cellular behaviors. The recent discovery of the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated (Cas) system from prokaryotic adaptive immune system demonstrated unprecedented genome editing efficiency and programmability in sequence specific genome editing of mammalian cells. In this thesis, I utilized the CRISPR-Cas system to construct a combinatorial genetic perturbation platform that enables massively parallel high throughput screening of multiple gene elements. This technology platform allows systematically interrogation of higher-order interactions of genetic regulators. The later part of the work described the establishment of a genomically encoded cellular recorder with the ability to longitudinally track and record molecular events in live animals. This cellular recorder encodes cellular memory through the quantitative accumulation of targeted genomic mutations, that allows mapping of a dynamical set of gene regulatory events without the need for continuous cell imaging or destructive sampling. Together, we envision these sets of technology and tools will offer new insights into cellular process in disease and in health.