| The discovery of induced pluripotent stem cells (iPSCs) has transformed the research of stem cells and provided infinite possibilities in regenerative medicine. In classical Yamanaka protocol, somatic cells from various sources can be reprogrammed to iPSCs with forced expression of Oct4, Sox2, Klf4, and cMyc. Numerous other combinations of factors and various delivery methods have also been developed to optimize the efficiency and accustom to different applications. These iPSCs are useful for disease modeling, toxicology studies and cell therapy. However, the molecular mechanisms of this transformation remain largely unclear. The transition from somatic cells to iPSCs involved comprehensive changes on epigenetic level of the cells induced by reprogramming factors. Reprogramming factor Klf4 can physically interact with Oct4 and Sox2. These three transcription factors co-occupy promoters of many pluripotency related genes, such as endogenous Nanog and Oct4. The physical interactions depend on the C2H2 zinc fingers in Klf4. Abrogation of these interactions will lead to failure of reprogramming due to the inability of defective complexes in activating key downstream genes. These results suggest that direct interactions between reprogramming factors are essential for initiating key downstream genes. During reprogramming, nuclear architecture of the cells also experience dramatic changes. In pluripotent stem cells (PSCs), endogenous Oct4 loci interact with distant regions in cis and in trans. Many of these long range interactions are specific to PSCs. PSC-specific interchromosomal interactions are established prior to transcriptional activation of endogenous Oct4 during reprogramming. In PSCs, Oct4-colocalized domains are enriched in active genes and pluripotency factor binding. Transcription of Oct4 is facilitated when the Oct4 locus is co-localized with its interchromosomal partners. Finally, depletion or overexpression of Klf4 causes changes in interchromosomal interactions prior to loss of Oct4 transcription and PSC differentiation, suggesting that Klf4 regulates interchromosomal interactions independent of its role as a transcription factor. Together these results reveal two novel essential factors in facilitating reprogramming: physical interactions between reprogramming factors and nuclear architecture dynamics. |
