Extended Data Fig. 7: TASOR or MPP8 depletion reduced H3K9me3 eSPAN bias at HO L1s in HeLa cells.
From: Asymmetric distribution of parental H3K9me3 in S phase silences L1 elements
a. H3K9me3 density at HO L1s in WT and TASOR KO HeLa cells based on published CUT&RUN datasets35. Heatmaps (left) were sorted by the average H3K9me3 signals of each row in WT cells, with average density shown at the right. L1s were separated based on the effects TASOR KO on H3K9me3 levels. Of the 393 TASOR regulated H3K9me3 loci identified by Douse et al.35 using a cutoff of log2 fold-change < −1, we found that 119 HO L1s identified in this study were located at these loci and defined them as the Down group. All other HO L1s were grouped as No-diff. b. H3K9me3 density at HO L1s in WT, MPP8 KO and TASOR KO HeLa cells based on published ChIP-seq datasets29, with heatmaps (top, sorted as in a) and average density (bottom) shown. HO L1s were grouped as in a. c. Immunoblots to confirm the knockdown (KD) of MPP8 and TASOR in HeLa cells. Note that while sgRNAs targeting MPP8 or TASOR were used to generate these cells, cells were pooled after selection, instead of cloned. Therefore, MPP8 and TASOR were only depleted and labeled as KD in HeLa cells. Note that ORF1p were markedly up-regulated, while total H3K9me3 levels didn’t change to a detectable degree. n = 3. d. H3K9me3 CUT&Tag signals at HO L1s in WT, MPP8 KD and TASOR KD HeLa cells. The datasets were generated in this study and HO L1s were grouped as in a. e. H3K9me3 CUT&Tag signals at HO L1s separated based on L1 length in WT, MPP8 KD and TASOR KD HeLa cells. HO L1s longer and shorter than the medium length were grouped as long (n = 5,615) and short (n = 5,653), respectively. f. Average H3K9me3 eSPAN bias around all 2,809 replication origins in WT, MPP8 KD and TASOR KD HeLa cells. g, h. Box plots of H3K9me3 eSPAN bias at HO L1s in WT, MPP8 KD and TASOR KD HeLa cells. HO L1s (n = 11,268) were grouped based on the effects of MPP8/TASOR KD on H3K9me3 density defined in a (g), or based on L1 length, as defined in e (h). Box plots (g, h) show the median, 25% and 75% quartiles and minimal and maximal values with p values by two-sided Mann–Whitney–Wilcoxon tests, with Bonferroni correction for multiple comparisons. Each panel is a representative of at least two independent experiments. See Materials and Methods for more details. For gel source data, see Supplementary Fig. 1.
