Unknown,Transcriptomics,Genomics,Proteomics

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The RNA Pol II Elongation Factor Ell3 Marks Enhancers in ES Cells and Primes Future Gene Activation

ABSTRACT: Enhancers play a central role in precisely regulating the spatiotemporal expression of developmentally regulated genes. The molecular mechanisms and factors required for controlling the inactive or poised enhancers and their function in future gene activation remain elusive. Here, we have identified that Ell3, a member of the Ell (Eleven-nineteen Lysine-rich Leukemia gene) family of RNA Pol II elongation factors, predominately occupies active and inactive enhancers of many developmentally regulated genes in mouse embryonic stem cells. Ell3 localizes to active enhancers of key stem cell renewal genes, but is not required for maintaining the undifferentiated state. Instead, Ell3 binding to inactive or poised enhancers is essential for marking them for stem cell specification through regulating the activation of key lineage-specific genes throughout differentiation. Ell3M-bM-^@M-^Ys association with enhancers is required for setting up proper Pol II occupancy at the proximal promoter regions of neighboring genes. This functional interaction between Ell3 and proximal promoter Pol II is dependent on cohesin. The depletion of cohesin subunits removes Ell3 from the enhancers and results in reduction in the levels of promoter proximally paused Pol II. Our study demonstrates that Ell3 is required for stem cell fate specification through regulating Pol II pausing on key developmental genes in the undifferentiated state and for their proper expression during differentiation. ChIP-seq of Ell3 in mES cells. ChIP-seq of Pol II in mES cells after Ell3 shRNA and non-targeting shRNA. RNA-seq of mES cells after Ell3 shRNA and non-targeting shRNA. RNA-seq of mouse embryoid bodies at day five after non-targeting shRNA.

ORGANISM(S): Mus musculus

SUBMITTER: Alexander Garruss

PROVIDER: E-GEOD-38148 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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The RNA Pol II elongation factor Ell3 marks enhancers in ES cells and primes future gene activation.

Lin Chengqi C Garruss Alexander S AS Luo Zhuojuan Z Guo Fengli F Shilatifard Ali A

Cell 20121227 1-2

Enhancers play a central role in precisely regulating the expression of developmentally regulated genes. However, the machineries required for enhancer-promoter communication have remained largely unknown. We have found that Ell3, a member of the Ell (eleven-nineteen lysine-rich leukemia gene) family of RNA Pol II elongation factors, occupies enhancers in embryonic stem cells. Ell3's association with enhancers is required for setting up proper Pol II occupancy at the promoter-proximal regions of ...[more]

PMID: 23273992

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Project description:Estrogen-related receptor M-NM-3 (ERRM-NM-3) serves a critical O2-dependent regulatory role in differentiation of human cytotrophoblasts to syncytiotrophoblast. In this study, we investigated expression of genes encoding tissue kallikreins (KLK1) and voltage-gated K+ channels (KV7) during differentiation of human trophoblasts in culture and the roles of ERRM-NM-3 and O2 tension in their regulation. Expression of KLK1 and the KV7 channel subunits, KCNQ1, KCNE1, KCNE3, KCNE5, increased during differentiation of cultured human trophoblast cells in a 20% O2 environment. Notably, together with ERRM-NM-3, expression of KLK1, KCNQ1, KCNE1, KCNE3 and KCNE5 was markedly reduced when cells were cultured in a hypoxic environment (2% O2). Moreover, upon transduction of trophoblast cells with shRNAs for endogenous ERRM-NM-3, KLK1, KCNQ1, KCNE1 and KCNE3 expression was significantly decreased. Promoter and site-directed mutagenesis studies in transfected cells identified putative ERRM-NM-3 response elements (ERREs) within the KLK1 and KCNE1 5'-flanking regions required for ERRM-NM-3 -stimulated transcriptional activity. Binding of endogenous ERRM-NM-3 to these ERREs increased during trophoblast differentiation in culture and was inhibited by hypoxia. The KV7 blocker linopirdine reduced hCG secretion and aggregation of cultured human trophoblasts, suggesting a possible role of KV7 channels in cell fusion and differentiation. Illumina gene expression arrays of cultured human trophoblast cells revealed several genes upregulated during syncytiotrophoblast differentiation and downregulated upon ERRM-NM-3 knockdown involved in cell differentiation, adhesion, and synthesis of steroid and peptide hormones required for placental development and function. Collectively, these findings suggest that ERRM-NM-3 mediates O2-dependent expression of genes involved in human trophoblast differentiation, function and vascular homeostasis. Illumina whole genome gene expression array analysis was performed on human trophoblasts before culture, or 72 h after infection with recombinant lentiviruses expressing ERRM-NM-3 shRNA, or a control, nonsilencing shRNA. RNA from three independent experiments using placental tissues from three abortuses was extracted by miRNeasyM-BM-. Mini Kit (Qiagen, Maryland, USA). RNA was labeled and hybridized to an Illumina HumanHT-12 v4 BeadChip, according to the manufacturerM-bM-^@M-^Ys protocol.

Dataset Information

The RNA Pol II Elongation Factor Ell3 Marks Enhancers in ES Cells and Primes Future Gene Activation

Publications

The RNA Pol II elongation factor Ell3 marks enhancers in ES cells and primes future gene activation.

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