Protein Arginine Methyltransferase 6 Regulates Embryonic Stem Cell Identity
1 Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore , Singapore, Singapore .Stem cells and development (Impact Factor: 3.73). 03/2012; 21(14):2613-22. DOI: 10.1089/scd.2011.0330
Histone arginine methylation has emerged as an important histone modification involved in gene regulation. Protein arginine methyltransferase (PRMT) 4 and 5 have been shown to play essential roles in early embryonic development and in embryonic stem (ES) cells. Recently, it has been reported that PRMT6-mediated di-methylation of histone H3 at arginine 2 (H3R2me2) can antagonize tri-methylation of histone H3 at lysine 4 (H3K4me3), which marks active genes. However, whether PRMT6 and PRMT6-mediated H3R2me2 play crucial roles in early embryonic development and ES cell identity remain unclear. Here, we have investigated their roles using gain and loss of function studies with mouse ES cells as a model system. We report that Prmt6 and histone H3R2 methylation levels increased when ES cells are induced to differentiate. Consistently, we find that differentiation of ES cells upon upregulation of Prmt6 is associated with decreased expression of pluripotency genes and increased expression of differentiation markers. We also observe that elevation of Prmt6 increases the methylation level of histone H3R2 and decreases H3K4me, Chd1, and Wdr5 levels at the promoter regions of Oct4 and Nanog. Surprisingly, knockdown of Prmt6 also leads to downregulation of pluripotency genes and induction of expression of differentiation markers suggesting that Prmt6 is important for ES cell pluripotency and self-renewal. Our results indicate that a critical level of Prmt6 and histone H3R2me must be maintained in mouse ES cells to sustain their pluripotency.
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ABSTRACT: Embryonic stem cells derived from the inner cell mass of blastocysts are pluripotent. Pluripotency is maintained by a transcriptional network in which Oct4 and Nanog are master regulators. Notably, several zinc finger transcription factors have important roles in this network. Patz1, a BTB/POZ domain-containing zinc finger protein, is expressed at higher levels in the inner cell mass relative to the trophectoderm. However, its function in pluripotency has been poorly studied. Here, we show that Patz1 is an important regulator of pluripotency in embryonic stem cells. Patz1 RNAi, chromatin immunoprecipitation and reporter assays indicate that Patz1 directly regulates Pou5f1, Nanog. Global transcriptome changes upon Patz1 knock-down largely involve up-regulation of apoptotic genes and down-regulation of cell cycle and cellular metabolism genes. Patz1 ChIP-sequencing further identified more than 5,000 binding sites of Patz1 in mouse genome, from which two binding motifs were extracted. Furthermore, Gene Ontology analysis of genes associated with the binding sites displays enrichment for proximity to developmental genes. In addition, embryoid body assays suggest that Patz1 represses developmental genes. Together, these results propose that Patz1 is important for embryonic stem cell pluripotency.Stem cells and development 12/2013; 23(10). DOI:10.1089/scd.2013.0430 · 3.73 Impact Factor
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ABSTRACT: Embryonic stem (ES) cells derived from the inner cell mass (ICM) of blastocysts are characterised by their ability to self-renew and their potential to differentiate into many different cell types. Recent studies have shown that zinc finger proteins are crucial for maintaining pluripotent ES cells. Mouse zinc finger protein 322a (Zfp322a) is expressed in the ICM of early mouse embryos. However, little is known regarding the role of Zfp322a in the pluripotency maintenance of mouse ES cells. Here, we report that Zfp322a is required for mES cell identity since depletion of Zfp322a directs mES cells towards differentiation. Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays revealed that Zfp322a binds to Pou5f1 and Nanog promoters and regulates their transcription. These data along with the results obtained from our ChIP-seq experiment showed that Zfp322a is an essential component of mES cell transcription regulatory network. Targets which are directly regulated by Zfp322a were identified by correlating the gene expression profile of Zfp322a RNAi-treated mES cells with the ChIP-seq results. These experiments revealed that Zfp322a inhibits mES cell differentiation by suppressing MAPK pathway. Additionally, Zfp322a is found to be a novel reprogramming factor that can replace Sox2 in the classical Yamanaka's factors (OSKM). It can be even used in combination with Yamanaka's factors and that addition leads to a higher reprogramming efficiency and to acceleration of the onset of the reprogramming process. Together, our results demonstrate that Zfp322a is a novel essential component of the transcription factor network which maintains the identity of mouse ES cells.PLoS Genetics 02/2014; 10(2):e1004038. DOI:10.1371/journal.pgen.1004038 · 7.53 Impact Factor
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ABSTRACT: Retinoids are morphogens and have been implicated in cell fate commitment of embryonic stem cells (ESCs) to neurons. Their effects are mediated by RAR and RXR nuclear receptors. However, transcriptional co-factors required for cell and gene-specific retinoid signaling are not known. Here we show that Protein aRginine Methyl Transferase (PRMT) 1 and 8 have key roles in determining retinoid regulated gene expression and cellular specification in a multistage neuronal differentiation of murine ESCs. PRMT1 acts as a selective modulator, providing the cells with a mechanism to reduce the potency of retinoid signals on regulatory "hotspots". PRMT8 is a retinoid receptor target gene itself and acts as a cell type specific transcriptional co-activator of retinoid signaling at later stages of differentiation. Lack of either of them leads to reduced nuclear arginine methylation, dysregulated neuronal gene expression and altered neuronal activity. Importantly, depletion of PRMT8 results in altered expression of a distinct set of genes, including markers of gliomagenesis. PRMT8 is almost entirely absent in human glioblastoma tissues. We propose that PRMT1 and PRMT8 serve as a rheostat of retinoid signaling to determine neuronal cell specification in a context-dependent manner, and might also be relevant in the development of human brain malignancy. Stem Cells 2014.Stem Cells 03/2015; 33(3). DOI:10.1002/stem.1894 · 6.52 Impact Factor
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