A structural role for histone methyltransferase MET-2 represses transcription independent of H3K9me catalysis [RNA-seq]

Packaging genomic regions into silenced heterochromatin is critical to maintain organismal viability and tissue identity. Methylation of histone H3 on lysine 9 (H3K9me) marks heterochromatin. Here we show that in addition to catalyzing H3K9me, the C. elegans histone methyltransferase MET-2 (SETDB1-like) has a second non-catalytic function that can itself contribute to gene repression. We find that subnuclear concentrates, or foci, of MET-2 correlate with efficient HMT activity, yet foci composed of catalytic-deficient MET-2 are also able to mediate transcriptional silencing. Genetic ablation of met-2 or physiological disruption of MET-2 foci by heat stress results in loss of silencing coincident with an increase in histone acetylation. Restoration of catalytically deficient MET-2 foci mitigates H3K9 and H3K27 hyperacetylation, gene derepression, and infertility, demonstrating a structural role for foci of a conserved heterochromatic histone methyltransferase in gene silencing independent of its enzymatic activity. Overall design: Total RNAseq of RNA isolated from early C. elegans embryos (~1-300 cell stage) in N2 (Bristol), met-2(n4256) III, and met-2(gw1660) mutants cultured at 20°C, or exposed to 37°C for 1hr. Extraction of RNA was performed according to the WormBook protocol (Stiernagle, 2006). Total RNA was purified using RNeasy kit (QIAGEN 74104) including DNase treatment. Depletion of ribosomal RNA was done for 5 lg of total RNA with Ribo-Zero? Margnetic Gold Kit (Epicentre MRGZG12324) and further concentrated with RNA Clean & Concentrator kit (Zymo Research R1015) according to corresponding manufacturers instructions.