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. 2021 Jun 23;40(30):4919–4929. doi: 10.1038/s41388-021-01900-8

Fig. 5. NF-κB binds the promoter regions of SPRY4-IT1 and transcriptionally upregulates its expression.

Fig. 5

A The sequence logo of NF-κB/p65 was predicted by the JASPAR database. B Schematic of predicted binding sites (BS) between SPRY4-IT1 and NF-κB/p65. C Construction of luciferase reporter vectors comprising different truncation variants of SPRY4-IT1. Dual-luciferase reporter assays were performed by co-transfecting the full-length SPRY4-IT1 promoter (SPRY4-F) or truncated SPRY4-IT1 (SPRY-P1, SPRY4-P2) with NF-κB/p65 cDNA plasmid or empty vectors in 293 T cells. Relative luciferase activity was then assayed. D ChIP assay showing endogenous NF-κB/p65 bound to the SPRY4-IT1 promoter in HCT 116 cells. E RT-qPCR analysis of SPRY4-IT1 expression in MCF-7, HCT 116, and OVCAR-3 cells following the transient transfection of NF-κB/p65 cDNA. F, G RT-qPCR analysis of SPRY4-IT1 expression in MDA-MB-231, SK-OV-3, and SW620 cells treated with an NF-κB inhibitor (F) and siRNA (G) for 48 h. H SPRY4-IT1 and NF-κB/p65 expression scores were determined by analyzing correlations between these markers in colorectal, ovarian, and breast cancers.