Reprogramming to Pluripotency Using Designer TALE Transcription Factors Targeting Enhancers

Abstract

SummaryThe modular DNA recognition code of the transcription-activator-like effectors (TALEs) from plant pathogenic bacterial genus Xanthomonas provides a powerful genetic tool to create designer transcription factors (dTFs) targeting specific DNA sequences for manipulating gene expression. Previous studies have suggested critical roles of enhancers in gene regulation and reprogramming. Here, we report dTF activator targeting the distal enhancer of the Pou5f1 (Oct4) locus induces epigenetic changes, reactivates its expression, and substitutes exogenous OCT4 in reprogramming mouse embryonic fibroblast cells (MEFs) to induced pluripotent stem cells (iPSCs). Similarly, dTF activator targeting a Nanog enhancer activates Nanog expression and reprograms epiblast stem cells (EpiSCs) to iPSCs. Conversely, dTF repressors targeting the same genetic elements inhibit expression of these loci, and effectively block reprogramming. This study indicates that dTFs targeting specific enhancers can be used to study other biological processes such as transdifferentiation or directed differentiation of stem cells.

Genome-Editing Technologies: Principles and Applications

“…Facilitated by many of the insights gained from zinc-finger transcription factor technology, both TALEs and CRISPR-Cas9 have now further expanded the possibilities of engineered transcriptional activators and repressors. For example, TALEs and CRISPR-Cas9 have enabled rapid construction of custom genetic circuits and logic gates (Gaber et al 2014;Lebar et al 2014;Liu et al 2014b), complex gene regulation networks (Nielsen and Voigt 2014;Nissim et al 2014), and even facilitated cellular reprogramming (Gao et al 2013) and the differentiation of mouse embryonic fibroblasts to skeletal myocytes (Chakraborty et al 2014). dCas9 transcriptional effectors have even been used to efficiently mediate repression and activation of endogenous genes in Drosophila (Lin et al 2015b) and in plant cells (Piatek et al 2015).…”

Enabling functional genomics with genome engineering

“…For instance, transcriptional manipulation mediated by synthetic DBPs has been most well-characterized when targeted to within 300 base pairs of TSSs. However, directed modulation of distal regulatory elements, such as enhancers, has recently been shown to be possible, albeit with varying efficacy (Gao et al 2013(Gao et al , 2014Mendenhall et al 2013;Ji et al 2014;Frank et al 2015;Hilton et al 2015;Kearns et al 2015). Enhancers represent dynamic genomic regulatory modules with differential functionality during cell lineage specification, and perturbation of enhancer function has been strongly associated with disease (Heinz et al 2015;Roadmap Epigenomics Consortium et al 2015).…”

“…Selective and targeted writing or erasure of appropriate epigenetic modifications can establish the causality of respective marks in determining gene expression and may also define their relevance in organismal development and in cellular responses to distinct stimuli. Furthermore, targeted activation or suppression of regulatory loci involved in lineage specification or reprogramming could have enormous biotechnological utility (Gao et al 2013;Chakraborty et al 2014;Ji et al 2014;Chavez et al 2015). Additionally, programmed looping to connect distal genomic regions can serve to define the function of specific genomic contacts (Deng et al 2014).…”

To Crispr And Beyond: The Evolution Of Genome Editing In Stem Cells

“…TALENs were rapidly adopted by the field for genome modifications, and within 5 years of deciphering the recognition code of TALEs, they had overtaken ZFNs future science group To CRISPR & beyond: the evolution of genome editing in stem cells Perspective in popularity (based on number annual of publications, Figure 2). TALENs were used to engineer mouse and human stem cells for targeted developmental studies and disease modeling [52,[55][56][57][58][59][60], on one-cell mouse embryos to generate genomic deletions for knockout studies [61], and for reprogramming of mESCs into iPS cells [62]. TALENs-mediated gene correction has been demonstrated in fibroblasts derived from epidermolysis bullosa patients [63].…”