Extended Data Fig. 5: MEN1 perturbation caused repeat expression alteration in A549 cells, related to Fig. 2.
(a) The distribution of peak length called by CREAM and MACS2 respectively. (b) Upper panel: Barplot showing the number of upregulated repeats in each subfamily. Lower panel: Boxplot showing permutation Z-score for upregulated repeats in each subfamily. Red color indicates statistical significance. Horizontal lines in the box represent the upper quartile, median, and the lower quartile from the top to the bottom. The vertical extending lines mark the 5th to 95th percentile. (c) Donut showing the number of upregulated repeats within and outside of the CREAM peak regions. (d) Number of downregulated repeats within a given distance of 1,857 peaks with increased H3K4me3 and MLL binding. Background are randomly selected repeat regions that do not show differential expression upon MEN1 knockout. p-value was calculated by one sided paired t-test. (e-i) ChIP-qPCR analysis of MLL1 (e), H3K4me3 (f), MEN1 (g), UTX (h) and H3K27me3 (i) in A549 cells with and without knockout of MEN1. 3’UTR of LINE1-HS and LINE1-PA2 were selected for analysis. GAPDH promoter was used as control for normalization. Mean ± s.e.m of 2 biological replicates were shown (unpaired two-tailed Student's t-test). *: p value <0.05; **: p value <0.01; ***: p value <0.001. (j) Bar plot showing the number of inverted Alus and non-inverted Alus induced by MEN1 knockout. (k) Quantifications of immunofluorescence staining of dsRNA (left panel) and γ-H2AX (right panel). For dsRNA staining, Mean ± s.e.m of quantifications from 12 randomly picked fields were shown (unpaired two-tailed Student's t-test). *: p value < 0.05; **: p value < 0.01; ***: p value < 0.001. For γ-H2AX, Mean ± s.e.m of quantifications from 95, 67 and 42 randomly picked cells were shown for sgCtrl, sgMEN1-1, and sgMEN1-2 group respectively (unpaired two-tailed Student's t-test). ****: p value < 0.0001. (l) ISG gene expression levels in MEN1-low vs MEN1-high patient tumors in the TCGA LUAD cohort. (m) qRT-PCR showing the relative expression of representative ISGs in MEN1 knockout A549 cells relative to control cells. Housekeeping gene TBP was used as control.Mean ± s.e.m of 3 biological replicates were shown (unpaired two-tailed Student's t-test). ****: p value <0.0001.
