Dynamic regulation of replication timing during cell lineage decisions

Abstract

During embryogenesis, the fetal liver is one of the first organs to develop after the heart, relying heavily on signals from the adjacent cardiac mesoderm. As lineage specification proceeds during development and cells differentiate, genomic regions are segregated into early or late replicating domains, establishing a cell type-specific replication timing (RT) program. Decades of intensive research have uncovered a strong correlation between developmental changes in RT, genome organization, and the transcriptome. While global changes in RT have been identified during distinct differentiation pathways, lineage-specific changes during organ development remain unexplored. Identifying factors regulating changes in replication timing pertaining to cardiac and hepatic development would help better understand how replication timing is regulated during lineage decisions and help establish better models to study congenital defects. Here, we sought to study the developmental switches in RT, and the corresponding changes in the transcriptome, of hESC-derived hepatocyte-like cells and cardiomyocyte-like cells. We also developed a novel multi-omics approach for the parallel analysis of replication timing and gene expression (PARTAGE), as well as an optimized method for live single-cell subcloning of HepG2 cells to enable genome editing experiments and generation of mutant cell lines.