Multiple layers of gene expression regulation during murine embryonic stem cell differentiation induced by Nanog down regulation

Gene expression is regulated at several distinct steps: modulation of chromatin structure, transcription, RNA processing, and translation. We have explored the contribution of all four layers of gene expression regulations to the development of murine embryonic stem (ES) cells. ES cells were induced to differentiate by down regulating a single gene, Nanog. Dynamic changes in chromatin structure, RNA transcription, RNA level, and final protein product were recorded using global measurement techniques, including RNA microarray, ChIP-on-chip and iTRAQ based mass spectrometry. Different patterns of gene expression regulatory events were analyzed using bioinformatic and statistic tools. Positive correlations between different gene expression steps were observed. We have also observed a time delay for chromatin structure change to affect gene expression change. Comparison between RNA polymerase attachment and RNA level suggests a possible role of post transcription regulation. There is evidence for a multifaceted regulation of protein production. About 40% of the protein changes are not correlated with RNA changes during this ES cell differentiation process, suggesting regulation at the translational or post translational levels. Post transcription control may determine the change of about 22% of the proteins. Modulation of chromatin structure and transcription may each determine the change of about 16% of the proteins. We have also found that genes with the same function are co-regulated at the same gene expression control step. We have reported the epigenetic change during ES cell differentiation in terms of histone isoforms change at the protein level and genomic histone acetylation dynamics at individual gene promoters. Our data on the multiple layers of gene expression after Nanog down regulation also allows for a deeper understanding of the Nanog downstream genes and Nanog binding genes. It helps to decipher the gene expression events during early ES cell differentiation. This is the first concerted epigenome, transcriptome, and proteome dynamic analysis.