Integrating single-cell imaging and RNA sequencing datasets links differentiation and morphogenetic dynamics

Basic helix-loop-helix genes, particularly proneural genes, are well-described triggers of cell differentiation, yet information on their dynamics is limited, notably in human development. Researchers at the Max Planck Institute of Molecular Cell Biology and Genetics focus on Neurogenin 3 (NEUROG3), which is crucial for pancreatic endocrine lineage initiation. By monitoring both NEUROG3 gene expression and protein in single cells using a knockin dual reporter in 2D and 3D models of human pancreas development, the researchers show an approximately 2-fold slower expression of human NEUROG3 than that of the mouse. They observe heterogeneous peak levels of NEUROG3 expression and reveal through long-term live imaging that both low and high NEUROG3 peak levels can trigger differentiation into hormone-expressing cells. Based on fluorescence intensity, the researchers statistically integrate single-cell transcriptome with dynamic behaviors of live cells and propose a data-mapping methodology applicable to other contexts. Using this methodology, they identify a role for KLK12 in motility at the onset of NEUROG3 expression.