Research Article

Gene bivalency at Polycomb domains regulates cranial neural crest positional identity

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Science  31 Mar 2017:
Vol. 355, Issue 6332, eaal2913
DOI: 10.1126/science.aal2913

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The epigenetics of face-making

How is it that our earlobes are attached to our ears and not our chins? Diverse bits of facial structure are derived from migrating neural crest cells. The cells start out similar but end up building very different facial structures. Neural crest cells destined for one structure can be rerouted to develop others, however. Minoux et al. found that neural crest cells share prepatterned poised chromatin states that are established before the cells migrate and retained during migration. Different developmental programs are unlocked when the migrating cells near their final location and interact with local patterning signals.

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Structured Abstract

INTRODUCTION

Craniofacial morphogenesis involves the cranial neural crest (NC) cells, a vertebrate-specific multipotent cell population that provides most of the head skeletogenic mesenchyme. Cranial NC cells delaminate from different points along the developing neural tube and migrate into distinct facial and pharyngeal arch processes, where they give rise to distinctly shaped cartilage and bone elements, which in turn assemble into a harmonious face. How do distinct cranial NC cell subpopulations acquire their regional identity, allowing them to generate the specific subsets of craniofacial elements appropriate to their position? Premigratory NC cells that contribute to frontonasal, maxillary, or mandibular processes share similar patterning potential, because they can replace each other in building a whole craniofacial skeleton. Such plasticity is maintained during and after migration until subpopulation-specific transcriptional identity and positional patterning programs are established as a result of interactions with their local surrounding environment. We asked how chromatin regulation may allow cranial NC cells to maintain broad patterning competence through migration while being poised to respond to local cues and induce position-specific transcriptional subprograms.

RATIONALE

We used genome-wide RNA sequencing (RNA-seq), chromatin immunoprecipitation followed by sequencing (ChIP-seq), and assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) and integrated the information to propose a model to explain how cranial NC subpopulations maintain broad patterning competence through chromatin epigenetic regulation and how transcription factor–dependent responses to local cues can modify the chromatin pattern to establish unique subpopulation-specific transcriptional subprograms. To this aim, we microdissected the Hox-free frontonasal, maxillary, mandibular, and the Hox-expressing second pharyngeal arch processes of E10.5 mouse embryos. We isolated the NC cell subpopulations from each of these prominences by cell sorting and analyzed their transcriptional state, as well as the H3K27me3, H3K4me2, and H3K27Ac histone modification and chromatin accessibility profiles at promoters and enhancers. We then compared these data sets with the transcriptional, histone mark, and chromatin accessibility profiles of the Hox-free NC premigratory progenitors and of E10.5 frontonasal, maxillary, mandibular, and second pharyngeal arch NC cell subpopulations in which we conditionally inactivated the Polycomb H3K27 methyltransferase gene enhancer of zeste homolog 2 (Ezh2cKO mutants).

RESULTS

Early postmigratory NC subpopulations contributing to distinct mouse craniofacial structures displayed similar chromatin accessibility patterns yet differed transcriptionally. The differentially expressed genes (positional genes) displayed accessible and H3K27me3+/H3K4me2+ bivalent enhancers and promoters, and were embedded in large Ezh2-dependent Polycomb domains, in the NC cell subpopulations in which they were silenced, indicating transcriptional poising. These postmigratory chromatin domains of poised gene regulation were inherited from NC premigratory progenitors. At Polycomb domains, H3K27me3 antagonized H3K4me2 deposition, which was restricted to accessible promoter and enhancer elements, preventing ectopic activation at inappropriate positions.

DISCUSSION

Our findings explain how cranial NC cell plasticity is maintained through migration until postmigratory stages. We propose that an Ezh2-dependent poised chromatin organization underlies the positional plasticity of cranial premigratory NC cell progenitors. This chromatin prepattern is maintained through migration. In response to position-specific environmental signals encountered by the NC cells during or after their migration, the regulatory elements and promoters of positional genes switch from a poised to an active chromatin state, contributing to establish NC subpopulation–specific transcriptional identities. This work contributes novel insights into the epigenetic regulation of face morphogenesis.

Epigenetic regulation of cranial NC cell identity.

A Polycomb-dependent poised chromatin organization underlies the positional plasticity of cranial premigratory NC cell progenitors. This chromatin prepattern is maintained through migration. In response to local cues encountered by the NC cells during or after their migration, the regulatory elements (E) and promoters (P) of differentially expressed genes switch from a poised to an active chromatin state, establishing transcriptional identities specific to subpopulations of NC cells.

Abstract

The cranial neural crest cells are multipotent cells that provide head skeletogenic mesenchyme and are crucial for craniofacial patterning. We analyzed the chromatin landscapes of mouse cranial neural crest subpopulations in vivo. Early postmigratory subpopulations contributing to distinct mouse craniofacial structures displayed similar chromatin accessibility patterns yet differed transcriptionally. Accessible promoters and enhancers of differentially silenced genes carried H3K27me3/H3K4me2 bivalent chromatin marks embedded in large enhancer of zeste homolog 2–dependent Polycomb domains, indicating transcriptional poising. These postmigratory bivalent chromatin regions were already present in premigratory progenitors. At Polycomb domains, H3K27me3 antagonized H3K4me2 deposition, which was restricted to accessible sites. Thus, bivalent Polycomb domains provide a chromatin template for the regulation of cranial neural crest cell positional identity in vivo, contributing insights into the epigenetic regulation of face morphogenesis.

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