Dynamics of Cell Fate Decisions during Chemically Induced Multi‐Lineage Trans‐Differentiation at Single‐Cell Level

Cell trans-differentiation offers a powerful means to manipulate cell identities. By exposing cells to a combination of small molecules (SMs), cell trans-differentiation can be induced in a simple and cost-effective manner. However, a comprehensive atlas detailing chemical-induced cell trans-differentiation across multiple cell fates has yet to be established. In this study, the underlying mechanisms of trans-differentiation is investigated and constructed an in-depth single-cell atlas of this process. The time-course trajectory is demonstrated for trans-differentiation of mouse embryonic fibroblasts (MEFs) into multiple cell lineages including epithelial, neural, extraembryonic endoderm like (XEN-like) cells, and endothelial cells, when induced by SMs cocktail 6TCF (E616452, tranylcypromine, CHIR99021, and forskolin). These trans-differentiated cells closely resemble various somatic cell types in the fetus. It is found that trans-differentiation is marked by dynamic shifts in entropy and the cell cycle during cell fate transitions. A common intermediate feature is revealed characterized by high ribosomal gene expression. This study combines high-resolution landscape with comparative analyses of trans-differentiation dynamics, providing new insights into the complex mechanisms driving cell fate determination in vitro. Future study shall explore the applicability of the model in human cell trans-differentiation.