Activation of the Imprinted Dlk1-Dio3 Region Correlates with Pluripotency Levels of Mouse Stem Cells

Abstract

Low reprogramming efficiency and reduced pluripotency have been the two major obstacles in induced pluripotent stem (iPS) cell research. An effective and quick method to assess the pluripotency levels of iPS cells at early stages would significantly increase the success rate of iPS cell generation and promote its applications. We have identified a conserved imprinted region of the mouse genome, the Dlk1-Dio3 region, which was activated in fully pluripotent mouse stem cells but repressed in partially pluripotent cells. The degree of activation of this region was positively correlated with the pluripotency levels of stem cells. A mammalian conserved cluster of microRNAs encoded by this region exhibited significant expression differences between full and partial pluripotent stem cells. Several microRNAs from this cluster potentially target components of the polycomb repressive complex 2 (PRC2) and may form a feedback regulatory loop resulting in the expression of all genes and non-coding RNAs encoded by this region in full pluripotent stem cells. No other genomic regions were found to exhibit such clear expression changes between cell lines with different pluripotency levels; therefore, the Dlk1-Dio3 region may serve as a marker to identify fully pluripotent iPS or embryonic stem cells from partial pluripotent cells. These findings also provide a step forward toward understanding the operating mechanisms during reprogramming to produce iPS cells and can potentially promote the application of iPS cells in regenerative medicine and cancer therapy.

Concise Review: Induced Pluripotent Stem Cells Versus Embryonic Stem Cells: Close Enough or Yet Too Far Apart?

“…Epigenetically, the locus is fully methylated in many iPSC lines, while some lines have only one allele silenced, as is the case in ESC. Functionally, it seems that iPSC with the Dlk1-Dio3 locus fully silenced can not form tetraploid complementation animals, and chimerism is also significantly lower when doing blastocyst injections, when compared with ESC [10,30].…”

microRNA-29b is a novel mediator of Sox2 function in the regulation of somatic cell reprogramming

“…Studies from Stadtfeld et al [20] and Liu et al [18] showed that the imprinted Dlk1-Dio3 region is mostly silenced by DNA methylation in OSKM-induced iPSCs and that the activation of this imprinted region might actually correlate with the degree of iPSC pluripotency. To probe the potential effect of miR-29b on the activation of the Dlk1-Dio3 region, we analyzed the expression of the genes and miRNAs in the Dlk1-Dio3 region in both OSKM-derived iPSC lines and OSKM + miR-29b-derived iPSC lines.…”

“…Treatment of cells with 5-Aza-2'-deoxycytidine (AZA, a non-specific inhibitor of DNMTs) facilitates the transition from somatic cells to pluripotent stem cells [14][15][16], and DNA demethylation is required for the reactivation of epithelial genes in the MET process at the early stage of iPSC generation [17]. Recent studies have found that the activation of certain imprinted regions, such as the Dlk1-Dio3 locus, is correlated with the developmental potential of fully pluripotent iPSCs [18,19]. Aberrant DNA hypermethylation might be the major cause of the Dlk1-Dio3 region silencing that prevents cells from be-coming fully pluripotent iPSCs [18,20].…”

“…Recent studies have found that the activation of certain imprinted regions, such as the Dlk1-Dio3 locus, is correlated with the developmental potential of fully pluripotent iPSCs [18,19]. Aberrant DNA hypermethylation might be the major cause of the Dlk1-Dio3 region silencing that prevents cells from be-coming fully pluripotent iPSCs [18,20]. DNMTs have thus been considered barriers to reprogramming.…”

Crucial Role of C-Myc in the Generation of Induced Pluripotent Stem Cells

“…S7A) [17]. We also examined the expression of imprinting genes, as a close relationship between iPSC quality, and these genes were recently suggested [25,26]. Nevertheless, no significant reduction in the expression of imprinting genes between 3F-iPSCs, 4F-iPSCs, or control ES cell lines was observed except for the 2A-3F-4-iPSC clone in which the expression of the Gtl2, Rtl1, Rian, and Mirg genes was suppressed (Supporting Information Fig.…”