Genetic depletion of bromodomain proteins phenocopies treatment with I-BET151.

(A) Left, Chromatin immunoprecipitation and sequencing (ChIP-seq) of four endogenously HA-tagged trypanosome bromodomain proteins showing localization to sites of transcription initiation for three of them. The region shown is a 184 kb region from chromosome 7. Arrows represent direction of transcription for PTUs. Areas where the arrows diverge are sites of transcription initiation. (B) Left, the percent of all MACS-derived peaks lying within 100 bp of a divergent strand switch region (Transcription Start Site, TSS), convergent strand switch region (Transcription Termination Site, TTS), both (a peak that is within a 100 bp of both a TSS and a TTS) or neither, a peak that is >100 bp away from an TTS or TSS. Right, the percent of all transcription start sites within 100 bp of a peak are shown for Bdf1,3,4. No Bdf2 MACS-derived peaks were within the FDR kd (top) or Bdf2KO (bottom) to untreated control cells. DESeq was used to normalize counts, compute log2(fold change) of treated cells over untreated cells, and calculate p-adjusted values. Dots in red indicate genes with fold changes of > 2 and a p-adjusted value KO (top) or Bdf3kd cells (middle), or I-BET151-treated cells (bottom) following washout of IBET-151 or Dox, and subsequent exposure to a 24 h treatment with one of two different differentiation triggers: incubation at 27°C (middle) or treatment with cis-aconitate (CA) at 37°C (right) compared to a control without any trigger (left, No stimulus). (F) Flow cytometry measuring the amount of primary anti-VSG antibody remaining on the surface following a 5-min incubation at the indicated temperatures, washout, fixation, and staining with secondary antibody in Dox-treated Bdf2KO (top panels) and Bdf3kd cells (bottom panels) compared to controls. Numerical data for Fig 5 is in S5 Data, except wiggle plots shown in Fig 5A, which are in S6 Data.