Intrinsic cellular heterogeneity directs divergent epithelial‐to‐mesenchymal transition outcomes

Epithelial-to-mesenchymal transition (EMT) is a fundamental process in development and disease; however, its study is often complicated by the intrinsic heterogeneity of commonly used epithelial cell models. Here, we show that pre-existing heterogeneity contributes to divergent outcomes following transforming growth factor-beta 3 (TGFβ3) treatment. Flow cytometry, gene expression profiling and imaging revealed persistent variability in E-cadherin (E-cad) and epithelial cell adhesion molecule (EpCAM) expression, with differentially sorted subpopulations displaying distinct transcriptional responses. Remarkably, some single-cell-derived clones exhibited paradoxical CDH1 upregulation during EMT, coinciding with GRHL3 induction and reflecting context-specific regulatory plasticity. Additionally, forskolin-mediated cyclic adenosine monophosphate (cAMP) activation enhanced epithelial stability and modulated EMT dynamics by preserving membrane-localised E-cadherin without preventing transcriptional repression of CDH1. Comparison with an epithelial subclone model further demonstrated how baseline identity shapes EMT trajectories. Moreover, in vivo analysis using zebrafish xenografts revealed that intrinsic epithelial features were associated with differential dissemination potential, reinforcing the connection between cellular identity and metastatic behaviour. These findings underline the importance of accounting for inherent heterogeneity in EMT models and demonstrate how intrinsic regulatory states and extrinsic signals intersect to shape epithelial plasticity.