Global epigenetic changes during somatic cell reprogramming to iPS cells

Abstract

Embryonic stem cells (ESCs) exhibit unique chromatin features, including a permissive transcriptional program and an open, decondensed chromatin state. Induced pluripotent stem cells (iPSCs), which are very similar to ESCs, hold great promise for therapy and basic research. However, the mechanisms by which reprogramming occurs and the chromatin organization that underlies the reprogramming process are largely unknown. Here we characterize and compare the epigenetic landscapes of partially and fully reprogrammed iPSCs to mouse embryonic fibroblasts (MEFs) and ESCs, which serves as a standard for pluripotency. Using immunofluorescence and biochemical fractionations, we analyzed the levels and distribution of a battery of histone modifications (H3ac, H4ac, H4K5ac, H3K9ac, H3K27ac, H3K4me3, H3K36me2, H3K9me3, H3K27me3, and γH2AX), as well as HP1α and lamin A. We find that fully reprogrammed iPSCs are epigenetically identical to ESCs, and that partially reprogrammed iPSCs are closer to MEFs. Intriguingly, combining both time-course reprogramming experiments and data from the partially reprogrammed iPSCs, we find that heterochromatin reorganization precedes Nanog expression and active histone marking. Together, these data delineate the global epigenetic state of iPSCs in conjunction with their pluripotent state, and demonstrate that heterochromatin precedes euchromatin in reorganization during reprogramming.

Embryonic stem cell and induced pluripotent stem cell: an epigenetic perspective

“…It appears that the chromatin reorganization events take place in a coordinated and sequential manner. Rearrangement of the heterochromatin, characterized by the presence of histone H3 lysine 9 trimethylation (H3K9me3) and HP1, precedes the activation of Nanog, while enrichment of euchromatin marks occurs concurrently with Nanog activation [14]. Consistently, heterochromatin is rearranged and becomes dispersed when partially reprogrammed cells are converted to iPSCs by dual inhibition of MEK and GSK3 [13].…”

“…During iPSC generation, the somatic cell chromatin needs to be reorganized to an ESC-like state with loosely organized heterochromatin and abundant euchromatin modifications [13,14]. It appears that the chromatin reorganization events take place in a coordinated and sequential manner.…”

“…The permissive chromatin state may lead to the activation of the pluripotency network with the help of the introduced reprogramming factors. In fact, expression of the other pluripotency gene, Nanog, has been shown to take place upon radical chromatin reorganization, and after this the pluripotency is established [13,14].…”

“…The epigenetic barrier restricting the pluripotency establishment in this step is likely reflected in the global non-permissive chromatin configuration and/or the inherent non-permissive chromatin context at the pioneer pluripotency genes. The molecular natures of the epigenetic barriers probably include H3K9me3/2 [13,14,71], H3K27me3 [110], insufficient histone acetylation [25,49,51,52], hypermethylation and hypo-hydroxylmethylation on DNA [7,147], and the lack of proper chromatin remodeling activities [34,42]. At this point, many questions remain unresolved.…”

“…iPSCs have been shown to be highly similar to ESCs, in terms of transcription program, chromatin modification profiles [5][6][7][8][9][10][11][12] and global chromatin configuration [6,13,14]. Functionally, at least some of the iPSCs have the developmental potential equivalent to ESCs, as entirely iPSC-derived animals ("all-iPSC" mice) can be generated through tetraploid complementation [15][16][17].…”