Figure 5.

p53-activating Mithramycin A boosts miR-3184-5p expression, which lowers ERBB2 expression and attenuates viability and invasion of cervical cancer cell lines. (A) p53, p21 and ERBB2 protein expression in cervical cancer cultures incubated with MM or vehicle (DMSO) assessed via WB. GAPDH was used as the loading control. (B) miR-3184-5p expression in cervical cancer cultures incubated with MM or vehicle assessed via RT-qPCR. U6 was used as the housekeeping control. (C) p53 and ERBB2 protein expression in cervical cancer cultures transfected with a p53 overexpression plasmid or empty plasmid control assessed via WB. GAPDH was used as the loading control. (D) miR-3184-5p expression in cervical cancer cultures transfected with a p53 overexpression plasmid or empty plasmid control assessed via RT-qPCR. U6 was used as the housekeeping control. (E) Representative images of Transwell and sphere-formation assays in (E) HeLa and (F) SiHa cells, and quantitative analysis of viability, invasion and sphere-formation of cells treated with MM or vehicle. (G) Schematic diagram of the p53 activator MM rescuing miR-3184-5p expression, thereby suppressing ERBB2 transcription. This attenuates PIK3CA activity, which stimulates cervical cancer cell viability, invasion and sphere-formation. Data are expressed as the mean ± SEM (n=3). **P<0.01 analyzed via unpaired Student's t-test. MM, Mithramycin A; Ctrl, control; WB, western blotting; RT-qPCR, reverse transcription-quantitative PCR; miR, microRNA; vec, vector; ERBB2, Erb-B2 Receptor Tyrosine Kinase 2; p-, phosphorylated; PIK3CA, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit α.