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Zic2 is an enhancer-binding factor required for embryonic stem cell specification.

ABSTRACT: The Zinc-finger protein of the cerebellum 2 (Zic2) is one of the vertebrate homologs of the Drosophila pair-rule gene odd-paired (opa). Our molecular and biochemical studies demonstrate that Zic2 preferentially binds to transcriptional enhancers and is required for the regulation of gene expression in embryonic stem cells. Detailed genome-wide and molecular studies reveal that Zic2 can function with Mbd3/NuRD in regulating the chromatin state and transcriptional output of genes linked to differentiation. Zic2 is required for proper differentiation of embryonic stem cells (ESCs), similar to what has been previously reported for Mbd3/NuRD. Our study identifies Zic2 as a key factor in the execution of transcriptional fine-tuning with Mbd3/NuRD in ESCs through interactions with enhancers. Our study also points to the role of the Zic family of proteins as enhancer-specific binding factors functioning in development.

SUBMITTER: Luo Z

PROVIDER: S-EPMC5009765 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature

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Zic2 is an enhancer-binding factor required for embryonic stem cell specification.

Luo Zhuojuan Z Gao Xin X Lin Chengqi C Smith Edwin R ER Marshall Stacy A SA Swanson Selene K SK Florens Laurence L Washburn Michael P MP Shilatifard Ali A

Molecular cell 20150201 4

The Zinc-finger protein of the cerebellum 2 (Zic2) is one of the vertebrate homologs of the Drosophila pair-rule gene odd-paired (opa). Our molecular and biochemical studies demonstrate that Zic2 preferentially binds to transcriptional enhancers and is required for the regulation of gene expression in embryonic stem cells. Detailed genome-wide and molecular studies reveal that Zic2 can function with Mbd3/NuRD in regulating the chromatin state and transcriptional output of genes linked to differe ...[more]

PMID: 25699711

Similar Datasets

Zic2, an enhancer-binding factor required for embryonic stem cell specification

Project description:The Zinc finger protein of the cerebellum 2 (Zic2) is one of the vertebrate homologs of the Drosophila pair-rule gene odd-paired (opa). Our molecular and biochemical studies have demonstrated that Zic2 to preferentially bind to transcriptional enhancers and functions as a cofactor that interacts with the NuRD complex in ES cells. Detailed genome-wide studies demonstrate that Zic2 function with Mbd3/NuRD in regulating the chromatin state and transcriptional output of genes linked to differentiation. Zic2 is dispensable for the selfrenewal of ES cells but is required for proper differentiation, similar to what has been previously reported for Mbd3/NuRD. Our study identifies Zic2 as a key factor in the execution of the pluripotency program with Mbd3/NuRD in ES cells. ChIP-seq of Zic2, Chd4, Mbd3 and Zic3 in mES cells. ChIP-seq of H3K27me3, H3K27ac, H3K4me3, H3K4me1 and PolII in mES cells after Zic2 shRNA and non-targeting shRNA. RNA-seq of mES cells after Zic2, Zic3, Mbd3 and Mta2 shRNA and non-targeting shRNA.

2015-02-23 | E-GEOD-61188 | biostudies-arrayexpress

Zic2, an enhancer-binding factor required for embryonic stem cell specification

2015-02-23 | GSE61188 | GEO

Primordial germ cell specification from embryonic stem cells.

Project description:BackgroundPrimordial germ cell (PGC) specification is the first crucial step in germ line development. However, owing to significant challenges regarding the in vivo system, such as the complex cellular environment and potential problems with embryo manipulation, it is desirable to generate embryonic stem (ES) cells that are capable of overcoming these aforementioned limitations in order to provide a potential in vitro model to recapitulate the developmental processes in vivo.Methodology and principal findingsHere, we studied the detailed process of PGC specification from stella-GFP ES cells. We first observed the heterogeneous expression of stella in ES cells. However, neither Stella-positive ES cells nor Stella-negative ES cells shared a similar gene expression pattern with either PGCs or PGC precursors. Second, we derived PGCs from ES cells using two differentiation methods, namely the attachment culture technique and the embryoid body (EB) method. Compared with PGCs derived via the attachment culture technique, PGCs derived via the EB method that had undergone the sequential erasure of Peg3 followed by Igf2r resulted in a cell line in which the expression dynamics of T, Fgf8 and Sox17, in addition to the expression of the epiblast markers, were more similar to the in vivo expression, thus demonstrating that the process of PGC derivation was more faithfully recapitulated using the EB method. Furthermore, we developed an in vitro model of PGC specification in a completely chemically defined medium (CDM) that indicated that BMP4 and Wnt3a promoted PGC derivation, whereas BMP8b and activinA had no observable effect on PGC derivation.Conclusions and significanceThe in vitro model we have established can recapitulate the developmental processes in vivo and provides new insights into the mechanism of PGC specification.

| S-EPMC2602984 | biostudies-literature

Cell-cell interaction modulates neuroectodermal specification of embryonic stem cells.

Project description:The controlled differentiation of embryonic stem (ES) cells is of utmost interest to their clinical, biotechnological, and basic science use. Many investigators have combinatorially assessed the role of specific soluble factors and extracellular matrices in guiding ES cell fate, yet the interaction between neighboring cells in these heterogeneous cultures has been poorly defined due to a lack of conventional tools to specifically uncouple these variables. Herein, we explored the role of cell-cell interactions during neuroectodermal specification of ES cells using a microfabricated cell pair array. We tracked differentiation events in situ, using an ES cell line expressing green fluorescent protein (GFP) under the regulation of the Sox1 gene promoter, an early marker of neuroectodermal germ cell commitment in the adult forebrain. We observed that a previously specified Sox1-GFP+ cell could induce the specification of an undifferentiated ES cell. This induction was modulated by the two cells being in contact and was dependent on the age of previously specified cell prior to coculture. A screen of candidate cell adhesion molecules revealed that the expression of connexin (Cx)-43 correlated with the age-dependent effect of cell contact in cell pair experiments. ES cells deficient in Cx-43 showed aberrant neuroectodermal specification and lineage commitment, highlighting the importance of gap junctional signaling in the development of this germ layer. Moreover, this study demonstrates the integration of microscale culture techniques to explore the biology of ES cells and gain insight into relevant developmental processes otherwise undefined due to bulk culture methods.

| S-EPMC2448393 | biostudies-literature

Enhancer decommissioning by LSD1 during embryonic stem cell differentiation.

Project description:Transcription factors and chromatin modifiers are important in the programming and reprogramming of cellular states during development. Transcription factors bind to enhancer elements and recruit coactivators and chromatin-modifying enzymes to facilitate transcription initiation. During differentiation a subset of these enhancers must be silenced, but the mechanisms underlying enhancer silencing are poorly understood. Here we show that the histone demethylase lysine-specific demethylase 1 (LSD1; ref. 5), which demethylates histone H3 on Lys?4 or Lys?9 (H3K4/K9), is essential in decommissioning enhancers during the differentiation of mouse embryonic stem cells (ESCs). LSD1 occupies enhancers of active genes that are critical for control of the state of ESCs. However, LSD1 is not essential for the maintenance of ESC identity. Instead, ESCs lacking LSD1 activity fail to differentiate fully, and ESC-specific enhancers fail to undergo the histone demethylation events associated with differentiation. At active enhancers, LSD1 is a component of the NuRD (nucleosome remodelling and histone deacetylase) complex, which contains additional subunits that are necessary for ESC differentiation. We propose that the LSD1-NuRD complex decommissions enhancers of the pluripotency program during differentiation, which is essential for the complete shutdown of the ESC gene expression program and the transition to new cell states.

| S-EPMC4144424 | biostudies-literature

A flexible repertoire of transcription factor binding sites and a diversity threshold determines enhancer activity in embryonic stem cells.

Project description:Transcriptional enhancers are critical for development and phenotype evolution and are often mutated in disease contexts; however, even in well-studied cell types, the sequence code conferring enhancer activity remains unknown. To examine the enhancer regulatory code for pluripotent stem cells, we identified genomic regions with conserved binding of multiple transcription factors in mouse and human embryonic stem cells (ESCs). Examination of these regions revealed that they contain on average 12.6 conserved transcription factor binding site (TFBS) sequences. Enriched TFBSs are a diverse repertoire of 70 different sequences representing the binding sequences of both known and novel ESC regulators. Using a diverse set of TFBSs from this repertoire was sufficient to construct short synthetic enhancers with activity comparable to native enhancers. Site-directed mutagenesis of conserved TFBSs in endogenous enhancers or TFBS deletion from synthetic sequences revealed a requirement for 10 or more different TFBSs. Furthermore, specific TFBSs, including the POU5F1:SOX2 comotif, are dispensable, despite cobinding the POU5F1 (also known as OCT4), SOX2, and NANOG master regulators of pluripotency. These findings reveal that a TFBS sequence diversity threshold overrides the need for optimized regulatory grammar and individual TFBSs that recruit specific master regulators.

| S-EPMC8015845 | biostudies-literature

PAX6D Instructs Neural Retinal Specification from Human Embryonic Stem Cell-Derived Neuroectoderm

Project description:PAX6 is essential for neural retina (NR) and forebrain development but how PAX6 instructs NR versus forebrain specification remains unknown. We found that the paired-less PAX6, PAX6D, is expressed in NR cells during human eye development and along human embryonic stem cell (hESC) specification to retinal cells. hESCs deficient for PAX6D failed to enter NR specification. Induced expression of PAX6D but not PAX6A in a PAX6-null background restored the NR specification capacity. ChIP-Seq, confirmed by functional assays, revealed a set of retinal genes and non-retinal neural genes that are potential targets of PAX6D, including WNT8B. Inhibition of WNTs or knocking down of WNT8B restored the NR specification capacity of neuroepithelia with PAX6D knockout whereas activation of WNTs blocked NR specification even when PAX6D was induced. Thus, PAX6D specifies neuroepithelia to NR cells via regulation of WNT8B.

| S-SCDT-EMBOR-2020-50000V1 | biostudies-other

PAX6D instructs neural retinal specification from human embryonic stem cell-derived neuroectoderm.

Project description:PAX6 is essential for neural retina (NR) and forebrain development but how PAX6 instructs NR versus forebrain specification remains unknown. We found that the paired-less PAX6, PAX6D, is expressed in NR cells during human eye development and along human embryonic stem cell (hESC) specification to retinal cells. hESCs deficient for PAX6D failed to enter NR specification. Induced expression of PAX6D but not PAX6A in a PAX6-null background restored the NR specification capacity. ChIP-Seq, confirmed by functional assays, revealed a set of retinal genes and non-retinal neural genes that are potential targets of PAX6D, including WNT8B. Inhibition of WNTs or knocking down of WNT8B restored the NR specification capacity of neuroepithelia with PAX6D knockout, whereas activation of WNTs blocked NR specification even when PAX6D was induced. Thus, PAX6D specifies neuroepithelia to NR cells via the regulation of WNT8B.

| S-EPMC7507545 | biostudies-literature

IFI16 promotes human embryonic stem cell trilineage specification through interaction with p53.

Project description:Transcriptional regulation plays an essential role in the self-renewal and differentiation of human embryonic stem cells (hESCs). However, how external signals disrupt the self-renewal regulatory network and further drive hESC differentiation remains largely unknown. Here, we found the immune regulative protein, gamma-interferon-inducible protein 16 (IFI16) was involved in the regulation of both self-renewal and differentiation gene expression during hESC trilineage specification through interaction with p53. IFI16 expression levels were upregulated through JNK activation. IFI16 knockdown delayed the downregulation of self-renewal gene expression and suppressed the upregulation of differentiation gene expression, while IFI16 overexpression accelerated trilineage specification. Furthermore, IFI16 stabilized p53-binding in the genome through IFI16-p53 interaction and differentially regulated self-renewal and differentiation gene expression. Together, our results suggest a particular role of IFI16 in differential gene expression regulation during trilineage specification of hESCs in a manner that is dependent on the genome-wide profile of p53-binding directed by IFI16-p53 interaction.

| S-EPMC7596047 | biostudies-literature

A Sox2 distal enhancer cluster regulates embryonic stem cell differentiation potential.

Project description:The Sox2 transcription factor must be robustly transcribed in embryonic stem (ES) cells to maintain pluripotency. Two gene-proximal enhancers, Sox2 regulatory region 1 (SRR1) and SRR2, display activity in reporter assays, but deleting SRR1 has no effect on pluripotency. We identified and functionally validated the sequences required for Sox2 transcription based on a computational model that predicted transcriptional enhancer elements within 130 kb of Sox2. Our reporter assays revealed three novel enhancers--SRR18, SRR107, and SRR111--that, through the formation of chromatin loops, form a chromatin complex with the Sox2 promoter in ES cells. Using the CRISPR/Cas9 system and F1 ES cells (Mus musculus(129) × Mus castaneus), we generated heterozygous deletions of each enhancer region, revealing that only the distal cluster containing SRR107 and SRR111, located >100 kb downstream from Sox2, is required for cis-regulation of Sox2 in ES cells. Furthermore, homozygous deletion of this distal Sox2 control region (SCR) caused significant reduction in Sox2 mRNA and protein levels, loss of ES cell colony morphology, genome-wide changes in gene expression, and impaired neuroectodermal formation upon spontaneous differentiation to embryoid bodies. Together, these data identify a distal control region essential for Sox2 transcription in ES cells.

| S-EPMC4265674 | biostudies-literature

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