Research Article

Causal role for inheritance of H3K27me3 in maintaining the OFF state of a Drosophila HOX gene

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Science  07 Apr 2017:
Vol. 356, Issue 6333, eaai8236
DOI: 10.1126/science.aai8236

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DNA sequence and inherited gene silencing

Cell fate decisions require a gene's transcriptional status, whether on or off, to be stably and heritably maintained over multiple cell generations. For silenced genes, heterochromatin domains are associated with specific histone posttranslational modifications, and these histone marks are maintained during DNA replication and chromosome duplication (see the Perspective by De and Kassis). Laprell et al. show that parental methylated histone H3 lysine 27 (H3K27) nucleosomes in Drosophila are inherited in daughter cells after replication and can repress transcription, but that they are not sufficient to propagate the mark. Trimethylation of newly incorporated nucleosomes requires recruitment of the methyltransferase Polycomb repressive complex 2 (PRC2) to neighboring cis-regulatory DNA elements. Coleman and Struhl demonstrate that H3K27 trimethylated nucleosomes play a causal role in transmitting epigenetic memory at a Drosophila HOX gene through anchoring of PRC2 at the Polycomb response element binding site. Wang and Moazed examine fission yeast and show that both sequence-dependent and chromodomain sequence-independent mechanisms are required for stable epigenetic inheritance of histone modifications and the epigenetic maintenance of silencing. These studies highlight the crucial role of DNA binding for heritable gene silencing during growth and development.

Science, this issue p. 85, p. eaai8236, p. 88; see also p. 28

Structured Abstract

INTRODUCTION

Animal development depends on the capacity of cells to respond to transient spatial and temporal cues by establishing heritable states of gene expression that specify cell fate. A long-standing question is whether such epigenetic states can be maintained by the inheritance of cis-acting chromatin modifications. To answer this question, it is necessary to identify those modifications that play a causal role in epigenetic memory and investigate how they are maintained and propagated through multiple rounds of replication.

RATIONALE

Many animals are composed of diverse body segments governed by evolutionarily conserved HOX gene complexes. Early in embryogenesis, cells in each segment heritably activate some HOX genes while silencing others. This provides a paradigm of epigenetic memory in which transient cues establish permanent ON and OFF states of gene expression. In the OFF state, HOX gene chromatin is marked with trimethylation of lysine 27 of histone H3 (H3K27me3) that is catalyzed by Polycomb repressive complex 2 (PRC2). Here, we investigated whether H3K27me3 nucleosomes are the carriers of epigenetic memory of the OFF state. To this end, we generated a transgenic lacZ reporter of the Drosophila HOX gene Ultrabithorax (Ubx), which recapitulates Ubx regulation throughout development. Deposition of H3K27me3 at this reporter depends on the presence of a Polycomb response element (PRE), the cis-acting DNA to which PRC2 is targeted. We monitored both the OFF state and H3K27me3 after inducing excision of the PRE at different time points during development. Using this strategy, in combination with manipulation of PRC2 activity and H3K27 substrate availability, we assessed the capacity of PRC2 that is not recruited to the PRE to propagate H3K27me3 and investigated whether there is a causal relationship between H3K27me3 inheritance and maintenance of the OFF state.

RESULTS

Focusing on the developing wing imaginal disc where the transgene is normally OFF, we find that PRE excision results in a release from silencing that depends on the cell-division–coupled dilution of H3K27me3 and occurs after a stereotyped number of cell divisions. Further, we show that the rate of dilution depends on the residual capacity of PRC2 that is not recruited by the PRE to propagate the mark. Reducing this capacity increases the rate of dilution and leads to a corresponding decrease in the number of times a cell can divide while maintaining the OFF state. These findings establish a causal relationship between inheritance of the H3K27me3 mark and memory of the OFF state.

CONCLUSION

Our results demonstrate (i) that H3K27me3, once established at a repressed Drosophila HOX gene, remains heritably associated with that gene; (ii) that local inheritance of the mark transmits the memory of the OFF state from one cell generation to the next; and (iii) that long-term memory depends on efficient copying of the mark after each replication cycle, a process that requires recruitment of PRC2 by the PRE. Taken together, these findings establish H3K27me3 as a determinant of epigenetic memory and identify efficient propagation of the mark by PRE-associated PRC2 as a requirement for perpetuating the memory.

Epigenetic silencing of a HOX gene by inheritance of H3K27 trimethylation.

Inheritance of repressive H3K27me3 chromatin depends on the transmission of parental H3K27me3 nucleosomes to daughter DNAs and their capacity to template the H3K27 trimethylation of newly incorporated, naïve nucleosomes by PRE-associated PRC2. After PRE excision, residual “free” PRC2 can copy the mark, albeit inefficiently, resulting in replication-coupled serial dilution of H3K27me3 and release from silencing. The number of divisions required depends on the rate of dilution, establishing a causal relationship between inheritance of the mark and memory of the OFF state.

Abstract

Many eukaryotic cells can respond to transient environmental or developmental stimuli with heritable changes in gene expression that are associated with nucleosome modifications. However, it remains uncertain whether modified nucleosomes play a causal role in transmitting such epigenetic memories, as opposed to controlling or merely reflecting transcriptional states inherited by other means. Here, we provide in vivo evidence that H3K27 trimethylated nucleosomes, once established at a repressed Drosophila HOX gene, remain heritably associated with that gene and can carry the memory of the silenced state through multiple rounds of replication, even when the capacity to copy the H3K27me3 mark to newly incorporated nucleosomes is diminished or abolished. Hence, in this context, the inheritance of H3K27 trimethylation conveys epigenetic memory.

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