Construction of a dynamic gene regulatory network required for cellular reprogramming

The generation of induced Pluripotent Stem Cells (iPSCs) from somatic cells provides an excellent model to study mechanisms of transcription factor-induced global alterations of the genome and the epigenome. Here, we have investigated the early transcriptional events of cellular reprogramming triggered by the co-expression of OSKM (Oct4, Sox2, Klf4 and c-Myc) in Mouse Embryonic Fibroblasts (MEFs) and mouse Hepatocytes (mHeps) and identified a novel gene regulatory network composed of 9 Transcriptional Regulators, which are directly targeted by OSKM. Functional studies using single and double shRNA knock-downs of any of these TRs caused disruption of the network and dramatic reductions in reprogramming efficiency, indicating that this novel gene regulatory network is essential for the induction and establishment of pluripotency. We demonstrate that the stochastic co-expression of the 9TR network components occurring in a remarkably small number of cells approximates the percentage of reprogrammed cells and is a result of dynamic molecular events. Thus, the early binding patterns of OSKM and the subsequent probabilistic co-expression of pivotal TRs in subpopulations of cells steer the reconstruction of a gene regulatory network crucial for the generation of iPSCs.