Recording Mammalian Development, Regeneration, and Tumorigenesis at Single-cell Resolution

Abstract

Single cells are the basic building blocks of tissues and organs, shaping their phenotypes and functions. Understanding the temporal dynamics and cellular lineage relationships at the single-cell level during mammalian development, tissue regeneration, and tumorigenesis provides critical insights into fundamental biological processes. This dissertation presents a multimodal framework to record the timing and clonal history of cellular events in vivo, integrating synthetic or naturally occurring genetic changes, while incorporating cell state and lineage information. This approach revealed tissue-specific cell expansion timelines and timing of progenitor specification during gut development. Further analysis revealed unconventional developmental relationships in mouse embryonic development, such as a somite-derived erythroid progenitor population. In the intestinal epithelium, this framework led to the discovery of persister intestinal stem cells (pISCs), a novel cell population specified during early embryogenesis that contributes to the adult stem cell pool. pISCs, marked by Tob2, play a critical role in regeneration following epithelial damage by modulating TGF-β signaling, which supports damage-induced cellular reprogramming. Our experimental platform also enables high-throughput CRISPR-based genetic screens using knockout vector libraries, which provides a valuable resource for genotype-to-phenotype mapping based on single-cell gene expression. Through these experiments, we identified novel factors that affected gene editing efficacy. Additionally, clonal analysis of murine adenomas and multi-omic profiling of human polyps reveal polyclonal initiation in colonic precancers, challenging traditional monoclonal models of tumorigenesis and shedding light on clonal selection during the transition from precancer to cancer. Together, this work lays the foundation for in vivo multi-modal lineage recording, offering new perspectives on the origins and timing of development, regeneration, and tumorigenesis in mammalian systems. The findings have significant implications for cancer biology, developmental biology, tissue regeneration, and also other potential applications in biomedical research.