Wnt dose escalation during the exit from pluripotency identifies tranilast as a regulator of cardiac mesoderm

Wnt signaling is a critical determinant of cell lineage development. This study used Wnt dose-dependent induction programs to gain insights into molecular regulation of stem cell differentiation. We performed single-cell RNA sequencing of hiPSCs responding to a dose escalation protocol with Wnt agonist CHIR-99021 during the exit from pluripotency to identify cell types and genetic activity driven by Wnt stimulation. Results of activated gene sets and cell types were used to build a multiple regression model that predicts the efficiency of cardiomyocyte differentiation. Cross-referencing Wnt-associated gene expression profiles to the Connectivity Map database, we identified the small-molecule drug, tranilast. We found that tranilast synergistically activates Wnt signaling to promote cardiac lineage differentiation, which we validate by in vitro analysis of hiPSC differentiation and in vivo analysis of developing quail embryos. Our study provides an integrated workflow that links experimental datasets, prediction models, and small-molecule databases to identify drug-like compounds that control cell differentiation. [Display omitted] •Wnt dose controls differentiation of mesendoderm gene programs and cell types•Multiple regression model predicts cardiac differentiation efficiency•Tranilast enhances differentiation by promoting mesoderm and suppressing endoderm•Tranilast synergistically promotes cardiac differentiation in vivo and in vitro Wu et al. use scRNA-seq to evaluate how Wnt signaling dose affects derivation of cell types from pluripotency. They build a computational model to identify a gene set predictive of cardiomyocyte differentiation and identify a small-molecule Wnt regulator, tranilast, that promotes cardiac differentiation both in vivo and in vitro.