Fig. 3
RELAFUS1 binds on specific DNA regions through the unique DNA-binding motif. a Top three transcription factor (TF) binding motifs enriched within the RELAFUS1-HA peaks identified by the Multiple Em for Motif Elicitation (MEME) tool in 293T/tv-a cells. E-value, enrichment p value. b RCAS-GFP, RELA-HA, RELAFUS1-HA (FUS1-HA) and RELAFUS1−S486E-HA (4E-HA) vector expression in 293T/tv-a cells. c–e Relative Nanoluc reporter activity to the RELAFUS1-responsive element (RE) normalized to the Firefly luciferase activity and GFP cells in the 293T/tv-a cells (n = 6 or 8, in technical triplicate). MEME-1, 2 and 3 denote RELAFUS1-responsive reporters for top three (3×) or five (5×) tandem RELAFUS1-motif as shown in Additional file 4: Fig. S3D–F. f Top TF binding motif enriched within the RELAFUS1-HA peaks in mEPN cells. g, h Relative Nanoluc reporter activity to the top three tandem (3×) or top five single RELAFUS1-MEME-2 motifs normalized to the Firefly luciferase activity and GFP cells in 293T cells (n = 4 or 5, in technical triplicate). i Relative Nanoluc reporter activity to the NF-κB-RE normalized to the Firefly luciferase activity and GFP cells in 293T/tv-a cells (n = 8, in technical triplicate). Analysis was done using RM one-way ANOVA (c-e) or paired two-tailed t-test (g–i). *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. j RCAS vector expression in 293T cells. C11-HA, RCAS-C11orf95-HA k Relative NFKBIA mRNA expression in 293T cells. Data (mean ± SD) are displayed as the relative ratio to GFP cells (n = 3, in technical quadruplicate). Analysis was done using paired two-tailed t-test. *p < 0.05; **p < 0.01. l IκBα protein expression in 293T cells. See Methods for j–l