Fig. 4

Phosphatidylinositol-4-phosphate 5-kinase type 1 gamma (PIP5K1C) is a direct target of miR-4649-5p. A Scheme illustrating the identification of PIP5K1C as a putative target of miR-4649-5p by RNA-seq analysis of SUM159 miR-4649-5p mimic transfected cells and subsequent in silico target predictions. At the bottom, a target sequence between miR-4649-5p and PIP5K1C is presented. B The downregulation of PIP5K1C by the miR-4649-5p mimic was confirmed in three transiently transfected (48 h) and one stable TNBC cell line by RT-qPCR (n = 3; mean ± SD; *p ≤ 0.05). C The downregulation of PIP5K1C protein by the miR-4649-5p mimic was confirmed 48 h after transduction in three TNBC cell lines by Western Blotting. Densitometric quantification of the representative blot on the right, normalized to the housekeeper Cofilin, is presented on the left. D A dual luciferase reporter assay giving evidence for the direct binding of the miR-4649-5p mimic to a reporter construct carrying a wild type (wt) target sequence of the PIP5K1C 3' UTR which was compared to an empty control vector. Binding of the miRNA, and thus luciferase signal reduction, was abolished when the PIP5K1C sequence was mutated (mut) (n = 3; mean ± SD; **p ≤ 0.01)