Single-cell analysis of cardiogenesis reveals basis for organ level developmental defects

Organogenesis involves integration of myriad cell types, and dysregulation of cellular gene networks results in birth defects, affecting 5 per cent of live births. Congenital heart defects (CHD) are the most common malformations and result from disruption of discrete subsets of cardiac progenitor cells(1), yet the transcriptional changes in individual progenitors that lead to organ-level defects remain unknown. Here, we employed single-cell RNA sequencing (scRNA-seq) to interrogate early cardiac progenitor cells as they become specified during normal and abnormal cardiogenesis, revealing how dysregulation of specific cellular sub-populations has catastrophic consequences. A network-based computational method for scRNA-seq that predicts lineage-specifying transcription factors(2,3) identified Hand2 as a specifier of outflow tract cells but not right ventricular cells, despite failure of right ventricular formation in Hand2-null mice(4). Temporal single-cell transcriptome analysis of Hand2-null embryos revealed failure of outflow tract myocardium specification, whereas right ventricular myocardium was specified but failed to properly differentiate and migrate. Loss of Hand2 also led to dysregulation of retinoic acid signaling and disruption of anterior-posterior patterning of cardiac progenitors. This work reveals transcriptional determinants that specify fate and differentiation in individual cardiac progenitor cells, and exposes mechanisms of disrupted cardiac development at single-cell resolution, providing a framework to investigate congenital heart defects.