First critical repressive H3K27me3 marks in embryonic stem cells identified using designed protein inhibitor

Moody, J. D.; Levy, Shiri; Mathieu, Julie; Xing, Yalan; Kim, Woojin; Cheng, Dong; Tempel, W.; Robitaille, Aaron M.; Dang, Luke T.; Ferreccio, Amy; Detraux, Damien; Sidhu, Sonia; Zhu, Licheng; Carter, Lauren; Xu, Chao; Valensisi, Cristina; Wang, Yuliang; Hawkins, R. David; Min, Jinrong; Moon, Randall T.; Orkin, Stuart H.; Baker, David C.

doi:10.1073/pnas.1706907114

Cited by 44 publications

(69 citation statements)

References 36 publications

(29 reference statements)

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“…The JARID2 2iL-I-F hESC mutants showed reduced H3K27me3 epigenetic repressive marks and reduced stem cell marker signature, showing that JARID2 is required to maintain the stem cell 2iL-I-F pluripotency state. PRC2 is not required in earliest na€ ıve state in mouse or human but shows requirement in the 2iL-I-F state [20]. We show that while other PRC2 components are expressed in both stages, JARID2 is dramatically downregulated in the earliest, 5iLA but not in the 2iL-I-F state.…”

Section: Introductionmentioning

confidence: 68%

“…Since multiple pluripotent stages have been stabilized from early human development [10][11][12][13][14], it is critical to analyze if PRC2 components are required for stemness in these stages. Using a computationally designed new protein, EED binder, we recently showed that similar to mouse, the earliest na€ ıve 5iLA hESC state is PRC2-independent, however, PRC2 is required to maintain na€ ıve 2iL-I-F and primed hESC pluripotent states [20]. Therefore, PRC2 is not required in earliest na€ ıve state in either mouse or in human, but is required in later stages [20].…”

Section: Jarid2 Expression In Pluripotent Stagesmentioning

confidence: 99%

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Inducible CRISPR genome editing platform in naive human embryonic stem cells reveals JARID2 function in self-renewal

et al. 2018

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To easily edit the genome of naïve human embryonic stem cells (hESC), we introduced a dual cassette encoding an inducible Cas9 into the AAVS1 site of naïve hESC (iCas9). The iCas9 line retained karyotypic stability, expression of pluripotency markers, differentiation potential, and stability in 5iLA and EPS pluripotency conditions. The iCas9 line induced efficient homology-directed repair (HDR) and non-homologous end joining (NHEJ) based mutations through CRISPR-Cas9 system. We utilized the iCas9 line to study the epigenetic regulator, PRC2 in early human pluripotency. The PRC2 requirement distinguishes between early pluripotency stages, however, what regulates PRC2 activity in these stages is not understood. We show reduced H3K27me3 and pluripotency markers in JARID2 2iL-I-F hESC mutants, indicating JARID2 requirement in maintenance of hESC 2iL-I-F state. These data suggest that JARID2 regulates PRC2 in 2iL-I-F state and the lack of PRC2 function in 5iLA state may be due to lack of sufficient JARID2 protein.

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Section: Introductionmentioning

confidence: 68%

Section: Jarid2 Expression In Pluripotent Stagesmentioning

confidence: 99%

Inducible CRISPR genome editing platform in naive human embryonic stem cells reveals JARID2 function in self-renewal

et al. 2018

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“…Interestingly, preventing the redistribution of epigenetic marks during the transition to a naïve state is sufficient to block changes in chromatin compaction at several exemplar regions, thereby directly linking epigenome remodelling with aspects of genome organisation (McLaughlin et al). The transcriptome and cell state of mouse and human naïve PSCs are largely unaffected by experimentally disrupting Polycomb levels (Galonska et al, 2015;Moody et al, 2017;Riising et al, 2014;Shan et al, 2017). In contrast, primed PSCs are sensitive to the removal of Polycomb proteins (Collinson et al, 2016;Moody et al, 2017;Shan et al, 2017;Wang et al, 2018).…”

Section: Discussionmentioning

confidence: 99%

“…The transcriptome and cell state of mouse and human naïve PSCs are largely unaffected by experimentally disrupting Polycomb levels (Galonska et al, 2015;Moody et al, 2017;Riising et al, 2014;Shan et al, 2017). In contrast, primed PSCs are sensitive to the removal of Polycomb proteins (Collinson et al, 2016;Moody et al, 2017;Shan et al, 2017;Wang et al, 2018). These observations collectively imply that the stable transition from a naïve to a primed state of pluripotency requires the reconfiguration of DNA interactions to provide a coordinated set of 'poised' regulatory signals to control promoter priming.…”

Section: Discussionmentioning

confidence: 99%

Network analysis of promoter interactions reveals the hierarchical differences in genome organisation between human pluripotent states

Chovanec

Collier

Krueger

et al. 2019

Preprint

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SUMMARYA complex and poorly understood interplay between 3D genome organisation, transcription factors and chromatin state underpins cell identity. To gain a systems-level understanding of this interplay, we generated a high-resolution atlas of annotated chromatin interactions in naïve and primed human pluripotent stem cells and developed a network-graph approach to examine the atlas at multiple spatial scales. Investigating chromatin interactions as a network uncovered highly connected hubs that changed substantially in interaction frequency and in transcriptional co-regulation between pluripotent states.Small hubs frequently merged to form larger networks in primed cells, often linked by newly-formed Polycomb-associated interactions. Importantly, we identified state-specific differences in enhancer activity and interactivity that corresponded with widespread reconfiguration of transcription factor binding and target gene expression. These findings provide multilayered insights into the gene regulatory control of human pluripotency and our systems-based network approach could be applied broadly to uncover new principles of 3D genome organisation. RESULTS Promoter interaction mapping in naïve and primed human PSCsWe used PCHi-C to profile the global, high-resolution interactomes of 22,101 promoters in isogenic human naïve and primed PSCs. There was a strong concordance in pairwise interaction read counts between the biological replicates of the same cell type (r 2 >0.95; Figure S1A); therefore we merged replicates for all downstream analyses. PCHi-C data normalisation and signal detection using the CHiCAGO pipeline (Cairns et al., 2016) identified 75,091 significant cis-interactions between baited promoters and other genomic regions in naïve PSCs, and 83,782 in primed PSCs ( Figure S1B). Just under half of the interactions were common to both cell types (n=39,360). As expected, trans-interactions represented a small minority of promoter interactions (354 interactions). In both cell types, the majority of significant interactions were between the promoters of protein-coding genes and non-promoter genomic regions ( Figure S1B).Processed datasets from this large-scale resource are available through the Open Science Framework (https://osf.io/XXXXX) and Table S1, and raw sequencing reads have been deposited to the Gene Expression Omnibus (accession GSEXXXXXX). Network visualisation of promoter interactomesWe next developed a computational approach called Canvas (Chromosome architecture network visualisation at scales) to visualise high-resolution, capture-based DNA interaction data at a network scale.Network graphs were constructed where each node of the network represents an individual HindIII genomic fragment (average size, 4 kb) and each edge represents a significant interaction between nodes ( Figure 1A). We combined all significant interactions detected in naïve and primed PSCs to produce a single, unified network graph, which retains information about whether an interaction is shared or cell type-specific ( Figu...

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“…Whereas some naive-specific genes (such as DPPA3 and TBX3) are activated relatively early, others (such as DPPA5, KLF17 and ZFP57) are fully induced only upon subsequent passaging (Collier et al, 2017). Naive resetting of hPSCs also involves profound metabolic changes, including increased oxidative phosphorylation (Takashima et al, 2014;Sperber et al, 2015) and glycolytic metabolism (Gu et al, 2016), and confers tolerance to the removal or inhibition of PRC2 enzymes (Moody et al, 2017;Shan et al, 2017).…”

Section: Interconversions Between Distinct Human Pluripotent Statesmentioning

confidence: 99%

Mechanisms of gene regulation in human embryos and pluripotent stem cells

Theunissen

Jaenisch

2017

Development

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Pluripotent stem cells have broad utility in biomedical research and their molecular regulation has thus garnered substantial interest. While the principles that establish and regulate pluripotency have been well defined in the mouse, it has been difficult to extrapolate these insights to the human system due to species-specific differences and the distinct developmental identities of mouse versus human embryonic stem cells. In this Review, we examine genome-wide approaches to elucidate the regulatory principles of pluripotency in human embryos and stem cells, and highlight where differences exist in the regulation of pluripotency in mice and humans. We review recent insights into the nature of human pluripotent cells , obtained by the deep sequencing of pre-implantation embryos. We also present an integrated overview of the principal layers of global gene regulation in human pluripotent stem cells. Finally, we discuss the transcriptional and epigenomic remodeling events associated with cell fate transitions into and out of human pluripotency.

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First critical repressive H3K27me3 marks in embryonic stem cells identified using designed protein inhibitor

Cited by 44 publications

References 36 publications

Inducible CRISPR genome editing platform in naive human embryonic stem cells reveals JARID2 function in self-renewal

Inducible CRISPR genome editing platform in naive human embryonic stem cells reveals JARID2 function in self-renewal

Network analysis of promoter interactions reveals the hierarchical differences in genome organisation between human pluripotent states

Mechanisms of gene regulation in human embryos and pluripotent stem cells

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