Fig. 2: Single cell gene expression analysis of GSC-0827 cells after KAT5 inhibition in vitro.
From: KAT5 regulates neurodevelopmental states associated with G0-like populations in glioblastoma
a UMAP projection of scRNA-seq data for sgCD8 (control) and sgKAT5 cells in GSC-0827 cells 5 days post nucleofection. Filtering scheme and data quality assessment for this data is available in Supplementary Fig. 4. b The UMAP projection from a showing de novo clusters generated. Cell cycle state predictions using the ccSeurat and ccAF classifiers and RNA velocity analysis for this data from are available in Supplementary Fig. 5. Associated data files include: cluster-based gene expression analysis (Supplementary Data 2) and gene set enrichment for top 200 expressed and top 200 depleted genes for each cluster (Supplementary Data 3, 4, respectively). For control cells, there is one G0-like cluster (cluster 4) which showed some weak expression for p53-associated target genes, suggestive of a DNA damage-induced G0-like state, a common feature of cultured cells (Arora et al., 2017; Spencer et al., 2013) also to some degree Neural G0 genes (see text). This state likely represents the p27hi Edu- population observed in vitro in Supplementary Fig. 1b, which are capable of re-entering the cell cycle. c MKI67/Ki67 gene expression, which is only expressed in cycling cells, among cells and clusters from (b). d Gene Set Variation Analysis (GSVA) associated with clusters and cell cycle phases showing MCM pathway required for initiation of DNA replication in early S-phase. Cell cycle pathways are shown in Supplementary Fig. 5. e Cell cycle phase predictions using the ccAF classifier for scRNA-seq data from (a). f Analysis of CCNB1 gene expression in ccAF predicted cell cycle phases: Neural G0 contains fewest cells expressing CCNB1, while G2/M the most. g Heatmap of representative genes upregulated in scRNA-seq clusters from (b). h Violin plots of gene expression module scores for each cell from scRNA-seq data of sgCD8A and sgKAT5KO GSC-0827 cells. oRG: outer radial glia. OPC oligodendrocyte precursor cells. Each data point = single cell. KS test was used to test significance (p < 0.0001)(1323 CD8 KO control cells; 840 KAT5 KO cells). Genes contained in each module are available in Supplementary Data 5. i Western blot validation studies of gene expression changes associated with loss of KAT5 activity. Protein extracts from GSC-0827 C13 cells were used from Dox+ or Dox- (7 days) conditions. Western blots were performed at least twice with similar results (except nAURKA = 1, nS100B = 1, nKAT5 = 4). Source data with exact p values are provided with this paper as a Source Data file.
