Among the epigenetic aberrations of iPSCs

Among the epigenetic aberrations of iPSCs, DNA methylation is of particular importance. Previous studies showed that unique de novo differentially methylated (DMR) or hydroxymethylated regions (hDMR) are present in iPSCs compared with hESCs (Lister et al., 2011; Wang et al., 2013). Furthermore, the retention of the epigenetic memory of donor cell types via cell-type-specific methylation affects the differentiation potential of iPSCs (Kim et al., 2011). There are three major enzymes that mediate DNA methylation. De novo DNA methyltransferases (DNMT3A and DNMT3B) are responsible for transferring a methyl moiety from S-adenosyl-methionine to cytosine to make 5-methylcytosine (5mC). DNMT1 together with hemi-methylated DNA-binding protein UHRF1 maintain 5-mC during cell-cycle progression (Jones, 2012). DNA demethylation, on the other hand, is either passive or indirect in mammalian cells. It has been shown to be mediated by enzymes recruited during histamine receptors or nucleotide excision DNA repair responses, as well as by cytidine deaminases (Wu and Zhang, 2010). Ten-eleven translocation proteins (TET1, TET2, and TET3) belonging to the family of 2-oxoglutarate- and iron (II)-dependent dioxygenases were also identified as DNA demethylation proteins (Kriaucionis and Heintz, 2009; Tahiliani et al., 2009). TETs were shown to catalyze the oxidation of 5mC into 5-hydroxymethylcytosine (5hmC) (Kriaucionis and Heintz, 2009; Tahiliani et al., 2009). TETs further convert 5-hmC to formylcytosine (5fC) and carboxycytosine (5caC), which undergo base excision repair by thymine-DNA glycosylase (TDG) (Ito et al., 2011; Shen and Zhang, 2013). Whereas 5mC is enriched in promoter regions of silent genes, 5mC in the gene body is positively correlated with gene expression (Ball et al., 2009; Lister et al., 2009). In contrast, 5hmC in both the promoter and gene body is associated with promoting gene expression (Song et al., 2011). MicroRNAs, or miRNAs, are a family of small ∼22 nt RNAs that regulate gene expression at the mRNA or protein level, and with functional implications in a wide range of biological processes (Bartel, 2004). miRNAs are extensively studied for their cell- and tissue-specific roles in cancer where they are significant contributors to epigenetic landscaping (Croce, 2009). The function of miRNAs was also explored in the context of somatic cell reprogramming. It was found that the miRNA 290–295 cluster is highly expressed in ESCs (Marson et al., 2008), and could enhance reprogramming efficiency in combination with Oct4, Sox2, and Klf4 (Judson et al., 2009; Nakagawa et al., 2008). It was also shown that miRNA cluster 302–367 (Anokye-Danso et al., 2011), or the cocktail miR-200c, miR-302, and miR-369 (Miyoshi et al., 2011) alone, could successfully reprogram both human and mouse cells to pluripotency, although efficiency is low (Lu et al., 2012). A diverse number of miRNA targeting processes such as mesenchymal-epithelial transition, apoptosis, and senescence, have been characterized and shown to modulate reprogramming in combination with the classical transcription factors (Bao et al., 2013). The miR-29 family, comprising miR-29a, miR-29b1, and miR-29c, is aberrantly expressed in various cancers, plays a role in extracellular matrix (ECM) production and fibrosis, and has also been shown to target DNA methylation enzymes Dnmt3a and Dnmt3b (Fabbri et al., 2007; Roderburg et al., 2011; Suh et al., 2012; Yang et al., 2013). More recently, with the help of our collaborators and others, we have shown that miR-29a also targets the TET protein family and TDG that convert 5mC to 5hmC and C (Cheng et al., 2013; Yang et al., 2013). Furthermore, miR-29 levels are high in senescent cells (Martinez et al., 2011) and repressed in the presence of Myc (Chang et al., 2008). Downregulation of miR-29a also showed some improvement of reprogramming efficiency in mouse fibroblasts, but its role in human reprogramming remains unexplored (Yang et al., 2011).