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A unique chromatin signature uncovers early developmental enhancers in humans

Abstract

Cell-fate transitions involve the integration of genomic information encoded by regulatory elements, such as enhancers, with the cellular environment1,2. However, identification of genomic sequences that control human embryonic development represents a formidable challenge3. Here we show that in human embryonic stem cells (hESCs), unique chromatin signatures identify two distinct classes of genomic elements, both of which are marked by the presence of chromatin regulators p300 and BRG1, monomethylation of histone H3 at lysine 4 (H3K4me1), and low nucleosomal density. In addition, elements of the first class are distinguished by the acetylation of histone H3 at lysine 27 (H3K27ac), overlap with previously characterized hESC enhancers, and are located proximally to genes expressed in hESCs and the epiblast. In contrast, elements of the second class, which we term ‘poised enhancers’, are distinguished by the absence of H3K27ac, enrichment of histone H3 lysine 27 trimethylation (H3K27me3), and are linked to genes inactive in hESCs and instead are involved in orchestrating early steps in embryogenesis, such as gastrulation, mesoderm formation and neurulation. Consistent with the poised identity, during differentiation of hESCs to neuroepithelium, a neuroectoderm-specific subset of poised enhancers acquires a chromatin signature associated with active enhancers. When assayed in zebrafish embryos, poised enhancers are able to direct cell-type and stage-specific expression characteristic of their proximal developmental gene, even in the absence of sequence conservation in the fish genome. Our data demonstrate that early developmental enhancers are epigenetically pre-marked in hESCs and indicate an unappreciated role of H3K27me3 at distal regulatory elements. Moreover, the wealth of new regulatory sequences identified here provides an invaluable resource for studies and isolation of transient, rare cell populations representing early stages of human embryogenesis.

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Figure 1: Unique chromatin signatures distinguish two classes of regulatory elements in hESCs.
Figure 2: Functional and molecular characterization of class I and II elements.
Figure 3: A subset of class II elements acquires active enhancer chromatin signature upon neuroectodermal differentiation.
Figure 4: Class II elements have developmental enhancer activity in vivo.

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Gene Expression Omnibus

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All sequencing data have been deposited in Gene Expression Omnibus (GEO) data repository under accession number GSE24447.

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Acknowledgements

We thank Wysocka laboratory members for ideas and manuscript comments; I. A. Shestopalov and J. K. Chen for sharing zebrafish resources, equipment and knowledge; T. Howes and D. M. Kingsley for the pT2HE vector; Z. Weng and A. Sidow for Illumina sequencing; and A. Valouev for discussion on ChIP-seq data analysis. This work was supported by WM Keck Foundation Distinguished Young Scholar in Biomedical Research Award and CIRM RN1 00579-1 grant to J.W. A.R.-I. was supported by an EMBO long-term fellowship.

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Authors and Affiliations

Authors

Contributions

A.R.-I. conceived the project, performed and interpreted most experiments, including all genomic data analyses. R.B. established hESC culture and differentiation and performed most zebrafish imaging. T.S. generated enhancer reporter constructs, and together with S.A.B. and A.R-I. participated in the in vivo enhancer screening. R.A.F. performed the RT-qPCR analysis of enhancer RNAs. J.W. contributed ideas and interpreted results. A.R-I. and J.W. wrote the manuscript with input from all authors.

Corresponding author

Correspondence to Joanna Wysocka.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-22 with legends and Supplementary Tables 1-4. (PDF 6621 kb)

Supplementary Data 1

This file contains genomic coordinates for all Class I, Class II and class II→I elements identified in this work. (XLS 472 kb)

Supplementary Data 2

This file contains the complete GREAT analysis list of overrepresented categories and statistical tests for Class I elements. (XLS 55 kb)

Supplementary Data 3

This file contains the complete GREAT analysis list of overrepresented categories and statistical tests for Class II elements. (XLS 61 kb)

Supplementary Data 4

This file contains the Complete GREAT analysis list of overrepresented categories and statistical tests for class II→I elements. (XLS 55 kb)

Supplementary Data 5

This file contains a list of primers used for ChIP-qPCR and RT-qPCR analysis. (XLS 57 kb)

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Rada-Iglesias, A., Bajpai, R., Swigut, T. et al. A unique chromatin signature uncovers early developmental enhancers in humans. Nature 470, 279–283 (2011). https://doi.org/10.1038/nature09692

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