Driver mutations in genes encoding histone H3 proteins resulting in p.Lys27Met substitutions (H3-K27M) are frequent in pediatric midline brain tumors. However, the precise mechanisms by which H3-K27M causes tumor initiation remain unclear. Here, we use human hindbrain neural stem cells to model the consequences of H3.3-K27M on the epigenomic landscape in a relevant developmental context. Genome-wide mapping of epitope-tagged histone H3.3 revealed that both the wild type and the K27M mutant incorporate abundantly at pre-existing active enhancers and promoters, and to a lesser extent at Polycomb repressive complex 2 (PRC2)-bound regions. At active enhancers, H3.3-K27M leads to focal H3K27ac loss, decreased chromatin accessibility and reduced transcriptional expression of nearby neurodevelopmental genes. In addition, H3.3-K27M deposition at a subset of PRC2 target genes leads to increased PRC2 and PRC1 binding and augmented transcriptional repression that can be partially reversed by PRC2 inhibitors. Our work suggests that, rather than imposing de novo transcriptional circuits, H3.3-K27M drives tumorigenesis by locking initiating cells in their pre-existing, immature epigenomic state, via disruption of PRC2 and enhancer functions.

编码组蛋白 H3 蛋白的基因驱动突变导致 p.Lys27Met 取代 (H3-K27M) 在儿科中线脑肿瘤中很常见。然而，H3-K27M 导致肿瘤发生的确切机制仍不清楚。在这里，我们使用人类后脑神经干细胞来模拟 H3.3-K27M 在相关发育背景下对表观基因组景观的影响。表位标记的组蛋白 H3.3 的全基因组图谱显示，野生型和 K27M 突变体都大量掺入预先存在的活性增强子和启动子中，并在较小程度上掺入 Polycomb 抑制复合物 2 (PRC2) 结合区域。在活性增强子处，H3.3-K27M 导致局灶性 H3K27ac 丢失、染色质可及性降低以及附近神经发育基因的转录表达减少。此外，H3.3-K27M 在 PRC2 靶基因子集上的沉积导致 PRC2 和 PRC1 结合增加，并增强转录抑制，而 PRC2 抑制剂可以部分逆转这种抑制。我们的工作表明，H3.3-K27M 不是强加从头转录回路，而是通过破坏 PRC2 和增强子功能，将起始细胞锁定在其预先存在的、不成熟的表观基因组状态，从而驱动肿瘤发生。