Fig. 1

Genetic Modification of Histone H3.3 expression in Pediatric Glioma Cells and Astrocytes. a Expression of RFP reporter was confirmed in > 95% of Doxycycline-induced cells. b H3.3G34V expression was significantly reduced in lentiviral-transduced KNS42 cells treated with Doxycycline, compared to no Doxycycline and control conditions. **p < 0.01, ****p < 0.0001. c H3.3G34V protein was expressed in lentiviral-transduced NHAs treated with doxycycline, with no H3.3G34V expression observed in controls. d Flow cytometry was used to select the top 30% cells with H3.3G34V knockdown and knock-in for subsequent ChIP-Seq analysis. e Genome browser view of H3.3G34V enrichment at the H3F3A locus across cell lines studied. Tracks highlighted in red are those conditions without H3.3G34V expression. f Metagene profile of H3.3G34V enrichment in the gene body across the top 1000 most H3.3G34V-enriched genes in KNS42. Reduction of H3.3G34V enrichment is observed following doxycycline-induced knockdown (light blue versus navy blue lines). Greater H3.3G34V enrichment is also observed in NHA transduced for H3.3G34V expression (green line) compared to control transduced for H3.3WT expression (orange line)