p300 compensates for the loss of BRD4 to rescue transcription after BET inhibition. (A-C) Tornado plots of BRD4 and p300 binding differences between BETi and DMSO conditions in KASUMI-1, SKNO1, and OCI-AML3 cells (24-hour treatment). Positive enrichment (in red) shows stronger binding upon BETi. Negative enrichment, colored in blue, shows loss of binding upon BETi. Shown are results from representative matched pairs (color-tri: red = positive enrichment upon BETi treatment; white = no change; petrol blue = negative enrichment upon BETi treatment). With the exception of BRD4 ChIP-Seq in KASUMI-1, which was performed once per condition, all experiments were performed with 2 biological replicates per condition. (D) Average binding curve profiles for p300 at the top 5% rescued sites in the indicated cell lines. Binding at the 5% best-scoring sites that were found in KASUMI1 (upper); binding profiles at the 5% best-scoring sites that were found in SKNO1 (middle); the same method for 5% best-scoring sites that were found in OCI-AML3 (lower). (E) Examples of BRD4 and p300 binding profiles in DMSO and BETi-treated KASUMI-1, SKNO1, and OCI-AML3 cells, to demonstrate the BETi-triggered increase of p300 binding at/near promoters of the main RUNX1 isoforms. (F) Bar plots of BRD4 (blue bars) and p300 (light orange bars) FCs in binding between the BETi and DMSO conditions in KASUMI-1 and SKNO1 cells (24-hour treatment) at the indicated MYC and RUNX1 regulatory regions. Shown are log2 FCs of ChIP-qPCR results from 3 biological replicates and SD. (G) Western blot for p300 and GAPDH in SKNO1 cells during a time lapse of 0 (DMSO-treated control), 2, 4, 12, and 24 hours of PROTAC-mediated p300 degradation via deg_p300. (H) Scheme of treatment with BETi for 24 hours and degradation of p300 within the last 4 hours of BETi. (I) Bar plots of p300 binding ratios to IgG in BETi and DMSO conditions with or without the addition of deg_p300 within the last 4 hours of treatment in SKNO1 cells. Shown are ChIP-qPCR results from 3 biological replicates at the RUNX1 promoter and 1 MYC enhancer and SD. (J) Analysis of qPCR expression of the indicated transcripts/genes in SKNO1 cells after treatment with either BETi (24 hours of treatment), deg_p300 (4 hours), or BETi (24 hours) with the addition of deg_p300 during the last 4 hours of treatment. Shown are log2 FCs normalized to DMSO-treated controls and SD from 3 biological replicates. (K) Volcano plots showing SLAM-Seq expression changes in SKNO1 cells after 24 hours of treatment with DMSO or BETi (same as supplemental Figure 1E) and DMSO or BETi plus the addition of deg_p300 within the last 4 hours of treatment. Three biological replicates were acquired per condition/time point. (L) SLAM-Seq intensity at the 933 rescued genes at 24 hours of BET inhibition (left) with the addition of deg_p300 during the last 4 hours of treatment, to be compared with the values at 0 hours (DMSO-treated controls), 4 hours, and 24 hours of BETi treatment in the background. Shown are log2 FC ratios to 0 hours (DMSO-treated controls). SLAM-Seq intensity at the indicated exemplar rescued genes at 24 hours of BET inhibition (right) with the addition of deg_p300 during the last 4 hours of treatment, to be compared with the values at 0 hours (DMSO), 4 hours, and 24 hours of BETi treatment in the background. Shown are log2 FC ratios to 0 hours (DMSO). Three biological replicates were acquired per condition and time point. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; IgG, immunoglobulin G.

p300 compensates for the loss of BRD4 to rescue transcription after BET inhibition. (A-C) Tornado plots of BRD4 and p300 binding differences between BETi and DMSO conditions in KASUMI-1, SKNO1, and OCI-AML3 cells (24-hour treatment). Positive enrichment (in red) shows stronger binding upon BETi. Negative enrichment, colored in blue, shows loss of binding upon BETi. Shown are results from representative matched pairs (color-tri: red = positive enrichment upon BETi treatment; white = no change; petrol blue = negative enrichment upon BETi treatment). With the exception of BRD4 ChIP-Seq in KASUMI-1, which was performed once per condition, all experiments were performed with 2 biological replicates per condition. (D) Average binding curve profiles for p300 at the top 5% rescued sites in the indicated cell lines. Binding at the 5% best-scoring sites that were found in KASUMI1 (upper); binding profiles at the 5% best-scoring sites that were found in SKNO1 (middle); the same method for 5% best-scoring sites that were found in OCI-AML3 (lower). (E) Examples of BRD4 and p300 binding profiles in DMSO and BETi-treated KASUMI-1, SKNO1, and OCI-AML3 cells, to demonstrate the BETi-triggered increase of p300 binding at/near promoters of the main RUNX1 isoforms. (F) Bar plots of BRD4 (blue bars) and p300 (light orange bars) FCs in binding between the BETi and DMSO conditions in KASUMI-1 and SKNO1 cells (24-hour treatment) at the indicated MYC and RUNX1 regulatory regions. Shown are log2 FCs of ChIP-qPCR results from 3 biological replicates and SD. (G) Western blot for p300 and GAPDH in SKNO1 cells during a time lapse of 0 (DMSO-treated control), 2, 4, 12, and 24 hours of PROTAC-mediated p300 degradation via deg_p300. (H) Scheme of treatment with BETi for 24 hours and degradation of p300 within the last 4 hours of BETi. (I) Bar plots of p300 binding ratios to IgG in BETi and DMSO conditions with or without the addition of deg_p300 within the last 4 hours of treatment in SKNO1 cells. Shown are ChIP-qPCR results from 3 biological replicates at the RUNX1 promoter and 1 MYC enhancer and SD. (J) Analysis of qPCR expression of the indicated transcripts/genes in SKNO1 cells after treatment with either BETi (24 hours of treatment), deg_p300 (4 hours), or BETi (24 hours) with the addition of deg_p300 during the last 4 hours of treatment. Shown are log2 FCs normalized to DMSO-treated controls and SD from 3 biological replicates. (K) Volcano plots showing SLAM-Seq expression changes in SKNO1 cells after 24 hours of treatment with DMSO or BETi (same as supplemental Figure 1E) and DMSO or BETi plus the addition of deg_p300 within the last 4 hours of treatment. Three biological replicates were acquired per condition/time point. (L) SLAM-Seq intensity at the 933 rescued genes at 24 hours of BET inhibition (left) with the addition of deg_p300 during the last 4 hours of treatment, to be compared with the values at 0 hours (DMSO-treated controls), 4 hours, and 24 hours of BETi treatment in the background. Shown are log2 FC ratios to 0 hours (DMSO-treated controls). SLAM-Seq intensity at the indicated exemplar rescued genes at 24 hours of BET inhibition (right) with the addition of deg_p300 during the last 4 hours of treatment, to be compared with the values at 0 hours (DMSO), 4 hours, and 24 hours of BETi treatment in the background. Shown are log2 FC ratios to 0 hours (DMSO). Three biological replicates were acquired per condition and time point. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; IgG, immunoglobulin G.

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