Sp5 induces the expression of Nanog to maintain mouse embryonic stem cell self-renewal

Tang, Ling; Wang, Manman; Liu, Dahai; Gong, Mengting; Ying, Qi‐Long; Ye, Shoudong

doi:10.1371/journal.pone.0185714

Cited by 12 publications

(10 citation statements)

References 43 publications

(42 reference statements)

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“…TFs that bind this expanded TFBS repertoire are likely to be important regulators in pluripotent cells which could drive more efficient reprogramming in combination with conventional reprogramming factors. In support of this, SP5, TFAP2C, GBX2, E2F3, ZIC3, FOXP1/FOXO1, ZFX and PRDM14 have recently been revealed as important regulators of pluripotency, self-renewal or reprogramming (Harel et al, 2012;Lim et al, 2007;Pastor et al, 2018;Seki, 2018;Tai and Ying, 2013;Tang et al, 2017Tang et al, , 2016Zhang et al, 2011a). Although examples exist where regulatory grammar is critical for activity as proposed by the enhanceosome model (Thanos and Maniatis, 1995), our findings indicate TFBS diversity above a threshold has a greater impact on enhancer activity.…”

Section: Discusionsupporting

confidence: 70%

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

Singh

Mullany

Moorthy

et al. 2020

Preprint

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Transcriptional enhancers are critical for development, phenotype evolution and often mutated in disease contexts; however, even in well-studied cell types, the sequence code conferring enhancer activity remains unknown. We found genomic regions with conserved binding of multiple transcription factors (TFs) in mouse and human embryonic stem cells (ESCs) are enriched in a diverse repertoire of transcription factor binding sites (TFBS) including known and novel ESC regulators. Remarkably, using a diverse set of TFBS from this repertoire was sufficient to construct short synthetic enhancers with activity comparable to native enhancers. Site directed mutagenesis of conserved TFBS in endogenous enhancers or TFBS deletion from synthetic sequences revealed a requirement for >10 different TFBS for robust activity. Specific TFBS, including the OCT4:SOX2 co-motif, are dispensable, despite co-binding the OCT4, SOX2 and NANOG master regulators of pluripotency. These findings reveal a TFBS diversity threshold overrides the need for optimized regulatory grammar and individual TFBS that bind specific master regulators. Highlights: Comparative epigenomics determines the enhancer sequence code and synthetic enhancer design. A diversity of >10 TFBS are required and sufficient for robust enhancer activity  >40 TFBS contribute to enhancer activity implicating new TFs in pluripotency maintenance  Increased TFBS diversity, above a threshold, overrides the need for regulatory grammar

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Section: Discusionsupporting

confidence: 70%

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

Singh

Mullany

Moorthy

et al. 2020

Preprint

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“…Both KLF4 and Sp5 have been shown to regulate naive pluripotency in part through the transcriptional regulation of Nanog , whose downregulation marks the exit of ESCs from the naive state into a state that is primed for lineage specification 29 – 31 . Interestingly, we observed downregulation of both Nanog and Klf4 in I-BRD9-treated ESCs (Fig.…”

Section: Resultsmentioning

confidence: 99%

A non-canonical BRD9-containing BAF chromatin remodeling complex regulates naive pluripotency in mouse embryonic stem cells

Gatchalian¹,

Malik²,

Ho³

et al. 2018

Nat Commun

175

203

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The role of individual subunits in the targeting and function of the mammalian BRG1-associated factors (BAF) complex in embryonic stem cell (ESC) pluripotency maintenance has not yet been elucidated. Here we find that the Bromodomain containing protein 9 (BRD9) and Glioma tumor suppressor candidate region gene 1 (GLTSCR1) or its paralog GLTSCR1-like (GLTSCR1L) define a smaller, non-canonical BAF complex (GBAF complex) in mouse ESCs that is distinct from the canonical ESC BAF complex (esBAF). GBAF and esBAF complexes are targeted to different genomic features, with GBAF co-localizing with key regulators of naive pluripotency, which is consistent with its specific function in maintaining naive pluripotency gene expression. BRD9 interacts with BRD4 in a bromodomain-dependent fashion, which leads to the recruitment of GBAF complexes to chromatin, explaining the functional similarity between these epigenetic regulators. Together, our results highlight the biological importance of BAF complex heterogeneity in maintaining the transcriptional network of pluripotency.

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“…SP5, a transcription factor, is expressed predominantly in gastrodermal epithelial stem cells and maintains selfrenewal of embryonic and neuromesodermal stem cells [50][51][52]. It is a target for WNT/β-catenin signaling and its regulation is maintained by WNT/β-catenin feedback loop [50].…”

Section: Discussionmentioning

confidence: 99%

G9a regulates tumorigenicity and stemness through genome-wide DNA methylation reprogramming in non-small cell lung cancer

et al. 2020

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Background: Eukaryotic histone methyltransferases 2 (EHMT2 or G9A) has been regarded as a potential target for non-small cell lung cancer (NSCLC) therapy. This study investigated the regulatory roles of G9A in tumorigenesis and stemness in NSCLC. We isolated and enriched tumor-initiating cells (TIC) from surgically resected NSCLC tissues by FACS and sphere formation assays. We then knocked down G9A using shRNA and carried out genome-wide 850K methylation array and RNA sequencing analyses. We carried out in vivo tumorigenecity asssay using mice xenografts and examined G9A interactions with its novel target using chromatin Immunoprecipitation (ChIP). Results: We identified 67 genes hypomethylated and 143 genes upregulated following G9A knockdown of which 43 genes were both hypomethylated and upregulated. We selected six genes (CDYL2, DPP4, SP5, FOXP1, STAMBPL1, and ROBO1) for validation. In addition, G9A expression was higher in TICs and targeting G9a by shRNA knockdown or by selective inhibitor UNC0642 significantly inhibited the expression of cancer stem cell markers and sphere forming capacity, in vitro proliferation, and in vivo growth. Further, transient overexpression of FOXP1, a protein may promote normal stem cell differentiation, in TICs resulted in downregulation of stem cell markers and sphere forming capacity and cell proliferation in vitro indicating that the genes we identified are directly regulated by G9A through aberrant DNA methylation and subsequent expression. Similarly, ChIP assay has shown that G9a interacts with its target genes through H3K9me2 and downregulation of H3K9me2 following G9a knockdown disrupts its interaction with its target genes. Conclusions: These data suggest that G9A is involved in lung cancer stemness through epigenetic mechanisms of maintaining DNA methylation of multiple lung cancer stem cell genes and their expression. Further, targeting G9A or its downstream genes could be a novel therapeutic approach in treating NSCLC patients.

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Sp5 induces the expression of Nanog to maintain mouse embryonic stem cell self-renewal

Cited by 12 publications

References 43 publications

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

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

A non-canonical BRD9-containing BAF chromatin remodeling complex regulates naive pluripotency in mouse embryonic stem cells

G9a regulates tumorigenicity and stemness through genome-wide DNA methylation reprogramming in non-small cell lung cancer

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