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. 2021 Jun 18;40(29):4759–4769. doi: 10.1038/s41388-021-01876-5

Fig. 6. The carboxyl-terminal amino acids flanking the FLI DNA-binding domain are essential for EWS/FLI-mediated oncogenic transformation.

Fig. 6

A Protein schematics of 3xFLAG-tagged (3 F) EWS/FLI constructs: EF, EF DBD and EF DBD+ are described in Fig. 2. EF DBD+ ΔN represents an EWS/FLI mutant where EWS is fused to the DBD+ version of FLI missing the 7amino-terminal amino acids to the DNA-binding domain; EF DBD+ ΔC represents an EWS/FLI mutant where EWS is fused to the DBD+ version of FLI missing the 10 carboxyl-terminal amino acids to the DNA-binding domain. B Western blot analysis of constructs expressed in A673 cells using our knock-down/rescue system. C Venn diagram overlap analysis of RNA-sequencing results (N = 3 biological replicates each). Overlap depicts genes called as significantly regulated by the listed construct compared to control cells (iEF + Empty Vector Cells). Genes were called as significantly regulated using an FDR cut-off of 0.05 (p-value of overlap < 2.2e−16). D Soft agar assay colony formation quantification of A673 knock-down/rescue cells containing the listed knock-down and rescue constructs. Data represented by mean ± SEM (N = 3 biological replicates with 2 technical replicates each). Asterisks indicate p-value < 0.05 as compared to negative control iEF + Empty Vector sample with no EWS/FLI expression.