H3K4 Methylation-Dependent Memory of Somatic Cell Identity Inhibits Reprogramming and Development of Nuclear Transfer Embryos

摘要

class="head no_bottom_margin" id="sec1title">IntroductionDuring development, cells lose their pluripotent status and acquire a stable cell identity, which only rarely, if ever, changes to another kind. Yet, somatic cells can be reprogrammed to another cell fate by nuclear transfer (NT) to eggs (), by the expression of a combination of transcription factors () or by cell-cell fusion (). In these reprogramming procedures, the gene-expression pattern and epigenetic state characteristic of one differentiated cell identity is erased and the gene expression pattern specific to another cell type is established.However, the efficiency of complete reprogramming via NT is low, as less than 10% of NT embryos generated from differentiated cells reach adulthood (, ). This led to the hypothesis that differentiated cells acquire a resistance to reprogramming procedures, which during normal development, helps to stabilize their cell fate. Due to this resistance, eggs cannot fully reprogram the incoming somatic nuclei, so that embryos with aberrant gene expression patterns arise and normal embryonic development is not supported (, , ). So far, it has been shown that a failure in reactivating genes, e.g., the pluripotency gene Oct4, during nuclear reprogramming is indicative of a poor developmental outcome of NT embryos (). Furthermore, epigenetic modifications inhibiting the re-activation of genes during the reprogramming procedure have been investigated and their removal has been utilized to improve reprogramming efficiency and to increase the viability of NT embryos (, , , , , ). However, the expression of donor cell-type-specific genes in the wrong cell type of NT embryos could also lead to a severe disruption of normal gene expression patterns resulting in developmental defects and embryonic lethality. Indeed, the existence of such an active transcription state memory has been suggested in NT and induced pluripotent stem cell (iPSC) experiments (href="#bib30" rid="bib30" class=" bibr popnode">Polo et al., 2010, href="#bib18" rid="bib18" class=" bibr popnode">Kim et al., 2011, href="#bib28" rid="bib28" class=" bibr popnode">Ng and Gurdon, 2005). Currently, however, the extent and functional importance of persistent donor-cell-type-specific gene expression in resistance to reprogramming is not known. Furthermore, the epigenetic mechanisms that confer memory of an active state of gene expression and that maintain the differentiated state of cells during nuclear reprogramming and embryonic development remain elusive.Here we show that in Xenopus and human NT embryos, memory of an active transcriptional state (ON-memory) is a phenomenon as widespread as the memory of an inactive transcriptional state. ON-memory genes are associated with increased levels of the active histone mark H3K4me3 when compared to properly reprogrammed genes in Xenopus and human somatic donor cells. Importantly, while a reduction in H3K4 methylation levels has little effect on gene expression in the donor cells, it significantly improves transcriptional reprogramming and enhances the developmental potential of the resultant NT embryos in Xenopus. Our study thus identifies H3K4 methylation as a critical epigenetic barrier in NT-mediated reprogramming and implicates its role as stabilization mechanism of cell differentiation.