The Pluripotency Regulator Zic3 Is a Direct Activator of the Nanog Promoter in ESCs

Abstract

The transcription factor Zic3 is required for maintenance of ESC pluripotency. By genome-wide chromatin immunoprecipitation (ChIP-chip) in ESCs, we have identified 379 direct Zic3 targets, many of which are functionally associated with pluripotency, cell cycle, proliferation, oncogenesis, and early embryogenesis. Through a computational analysis of Zic3 target sequences, we have identified a novel Zic3 consensus binding motif (5 0 -CC C / T GCTGGG-3 0 ). ChIP results and in vitro DNA binding assays revealed that Zic3 binds with high affinity and specificity on the Nanog promoter. Here, we demonstrate that Zic3 functions as a transcriptional activator of the Nanog promoter in three ways:(a) Nanog transcript levels are sustained with Zic3 overexpression in differentiating ESCs, (b) Zic3 depletion in ESCs downregulates Nanog promoter activity, and (c) Zic3 overexpression leads to increased Nanog promoter activity. Furthermore, the activity of a mutant Nanog promoter with ablated Oct4/Sox2 binding is rescued by Zic3 overexpression to nearly wild-type levels. This indicates that Nanog is a positive transcriptional target of Zic3 in a mechanism that is independent of Oct4/Sox2 binding. Hence, we demonstrate an important pathway for regulation of Nanog expression in pluripotent ESCs through direct activation by Zic3. STEM CELLS

Direct lineage conversion: induced neuronal cells and induced neural stem cells

“…Kumar et al demon-strated that the co-transduction of Zic3 with Oct4, Sox2, and Klf4 could convert human fibroblasts to iNS cells. Zic3, a member of the Gli superfamily, has been implicated in the maintenance of pluripotency in mouse and human embryonic stem cells (ESCs), where it directly controls the ex-pression of Nanog (Lim et al, 2007(Lim et al, , 2010. Zic3 is also an immediate early gene induced by fibroblast growth factor (FGF) signaling during neural specification (Marchal et al, 2009).…”

Changing Faces of Transcriptional Regulation Reflected by Zic3

“…Second, a consensus binding motif of Zic3 in zebrafish is highly similar to that found in mouse ES cells [25] (Hong et al , unpublished). In sharp contrast, most of the surrounding regions appear to be poorly conserved in evolution.…”

“…Profiling of Zic3-binding sites using ChIP-chip in mouse ES cells covered 17,000 promoters spanning regions between -5.5kb to +2.5kb of transcription start sites and revealed potential involvement of Zic3 in regulation of some 300 genes, including several linked with pluripotency [25]. Application of next generation sequencing (NGS) allows an unbiased genome-wide assessment of Zic3 binding sites by ChIP-seq, which revealed that a third of Zic3 binding sites were identified within the promoter region (Hong et al , unpublished).…”

“…Analysis of Zic3 interactions in mouse ES cells revealed a role of this TF in inhibiting endodermal differentiation [25, 39]. At the same time when co-expressed with the Oct4, Sox2, and Klf4, Zic3 enhances the reprogramming of human fibroblasts into cells that resemble neural progenitors [40].…”

Identificação de vias moduladas por microRNAs na diferenciação celular e manutenção da pluripotência em células humanas

“…Esse fenômeno pode ser explicado, por exemplo, pela propriedade autorregulatória de NANOG, que ao interagir com diferentes reguladores transcricionais, pode induzir ou reprimir a transcrição de seu próprio gene (FIDALGO et al, 2012;LIANG et al, 2008;LIM et al, 2010;LOH et al, 2006).…”