Structural basis for PRC2 decoding of active histone methylation marks H3K36me2/3.

Area of Science:

• Chromatin biology
• Epigenetics
• Molecular mechanisms of gene regulation

Background:

• Polycomb group (PcG) proteins are essential epigenetic regulators.
• PRC2 catalyzes histone H3 trimethylation at lysine 27 (H3K27me3), a mark associated with gene silencing.
• H3K36 methylation (H3K36me2/3) at active genes inhibits PRC2, but the mechanism remains unclear.

Purpose of the Study:

• To elucidate the structural basis for H3K36-mediated inhibition of PRC2.
• To understand how PRC2 interacts with nucleosomal DNA and histone tails.
• To investigate the functional consequences of H3K36 modifications on Polycomb repression.

Main Methods:

• Cryo-electron microscopy (cryo-EM) to determine the structure of PRC2 bound to dinucleosomes.
• Site-directed mutagenesis of histone H3 (H3K36A, H3K36R).
• In vitro biochemical assays to measure H3K27 methylation activity.
• Analysis of Polycomb repression of HOX genes in Drosophila H3K36 mutants.

Main Results:

• The cryo-EM structure reveals how PRC2's catalytic subunit EZH2 binds nucleosomal DNA, positioning the H3 N-terminus for H3K27 methylation.
• Unmodified H3K36 occupies a key site at the EZH2-DNA interface, suggesting a role in PRC2 regulation.
• Mutations of H3K36 to alanine or arginine impair PRC2's H3K27 methylation activity in vitro.
• Drosophila H3K36 mutants exhibit reduced H3K27me3 levels and defective Polycomb repression of HOX genes.

Conclusions:

• A network of interactions involving EZH2, nucleosomal DNA, and the H3 N-terminus dictates PRC2's substrate access and allosteric regulation.
• The structure explains how H3K36 methylation acts as an inhibitory signal in transcriptionally active chromatin.
• This mechanism ensures proper gene silencing by Polycomb proteins and prevents repression of active genes.