Fig. 3: DNA-binding and genomic localization properties of EWS/FLI unaltered by deletions flanking the FLI DNA-binding domain. | Oncogene

Fig. 3: DNA-binding and genomic localization properties of EWS/FLI unaltered by deletions flanking the FLI DNA-binding domain.

From: The FLI portion of EWS/FLI contributes a transcriptional regulatory function that is distinct and separable from its DNA-binding function in Ewing sarcoma

Fig. 3

A Protein schematic of FLI DBD and FLI DBD+ recombinant protein (with C-terminal 6xHistidine-tag [6xHis]). B–D Fluorescence anisotropy assay results for FLI DBD and FLI DBD+ recombinant proteins (0–20 μM) on 5 nM fluorescein-labeled DNA sequences: B ETS high-affinity (HA) site DNA, C 2x-repeat GGAA μSat DNA, and D 20x-repeat GGAA μSat DNA (N = 2 biological replicates, 3 technical replicates each). Dissociation constants (KD) for FLI DBD and FLI DBD+ are noted for each DNA response element. E Venn diagram comparing peaks called in CUT&RUN for EWS/FLI construct localization in knock-down/rescue cells (EF = iEF + EF; EF DBD = iEF + EF DBD; EF DBD + = iEF + EF DBD+) when compared to cells that did not contain a rescue construct (iEF + Empty Vector) (adjusted p-value (FDR) < 0.05; N = 2 biological replicates each). The number of peaks overlapping between constructs are indicated on the Venn diagram. F–H Representative CUT&RUN peak tracks from IGV are shown for iEF + Empty Vector (EF KD), EF, EF DBD, and EF DBD+ samples. Examples of peaks from EWS/FLI-associated HA-site regulated genes ([F] STEAP1 and [G] BIRC2) and GGAA-μSat-regulated genes ([H] GSTM4) are highlighted. Peak track scales are shown on the right.

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