Dependence of cell fate potential and cadherin switching on primitive streak coordinate during differentiation of human pluripotent stem cells
During gastrulation, the primitive streak (PS) forms and begins to differentiate into mesendodermal subtypes. This process involves an epithelial-mesenchymal transition (EMT),which is marked by cadherin switching, where E-Cadherin is downregulated, and N-Cadherin is upregulated.To understand the relationships between differentiation, EMT, and cadherin switching, we made measurements of these processes during differentiation of human pluripotent stem cells (hPSCs) to PS and subsequently to mesendoderm subtypes using established protocols, as well as variants in which signaling through key pathways including Activin, BMP, and Wnt were modulated.We found that perturbing signaling so that cells acquired identities ranging from anterior to posterior PS had little impact on the subsequent differentiation potential of cells but strongly impacted the degree of cadherin switching. The degree of E-Cadherin downregulation and N-Cadherin upregulation were uncorrelated and had different dependence on signaling. The exception to the broad potential of cells throughout the PS was the loss of definitive endoderm potential in cells with mid to posterior PS identities. Thus, cells induced to different PS coordinates had similar potential within the mesoderm but differed in cadherin switching. Consistently, E-Cadherin knockout or overexpression did not alter cell fates outcomes during differentiation. Overall, although all processes are regulated by the same set of signaling pathways, the extent of cadherin switching and EMT can vary substantially within cells adopting the same cell fate. Overall design: RNA-seq of human pluripotent stem cells (ESI017) directed for 24 h into four primitive streak populations - Definitive Endoderm(day1) (DE(a)), Anterior PS (APS), Mid PS (MPS), Posterior PS (PPS) - and of matched cultures further differentiated for an additional 24 h toward Definitive Endoderm(day2) (DE(b)), Paraxial Mesoderm (PM), or Lateral Mesoderm (LM). Each condition was collected in 2 biological replicates.