Lysine 27 of replication-independent histone H3.3 is required for Polycomb target gene silencing but not for gene activation

Modifications of histone H3 lysine 27 (H3K27) are strongly correlated with gene activation and gene repression. The histones bearing these modifications are encoded by both the replication-dependent H3 genes, and the replication-independent H3.3 genes. To examine the role that K27 modification of the H3.3 genes plays during development, we use CRISPR/Cas9 to mutate the endogenous H3.3 genes to a non-modifiable residue and performed RNAseq. We find that H3.3K27 is required for Polycomb target gene silencing, but this requirement is at a later stage of development than the requirement for replication-dependent H3K27. Notably, we find no evidence of transcriptional defects in H3.3K27 mutants, despite the strong correlation between H3.3K27 acetylation and active transcription. Overall design: Total-RNAseq from whole, male pharates in wildtype, H3.3B K27R single mutants, and H3.3A K27R; H3.3B K27R double mutants.