Empty vector control

Briefly, cells were seeded in six-well plates, co-transfected with miR-194 precursor vector (p-CMV-miR-194) or empty vector control (Origene, Rockville, MD, USA) and a wild-type AKT2 3′UTR reporter construct (pmiR-GLO-AKT2-3′UTR) or pmir-GLO-AKT2-3′UTR mutant or empty vector with the firefly luciferase reporter and the control vector Renilla luciferase, pRL-TK (Promega, Madison, WI, USA) using lipofectamine 2000 following the manufacturer's protocol. Renilla luciferase vector (pRL-TK) served as an internal control and was included in all samples. After 48 h, cells were lysed and Firefly and Renilla activities were measured sequentially using the dual luciferase assay kit (PRE1910, Promega). Luminescence was measured by a luminometer (Turner model 9100-002, Turner Biosystems Inc., Sunnyvale, CA, USA).

The miRNA mimic, miRNA inhibitor, STAT3 siRNA, and scrambled siRNA were synthesized by RiBoBio (China). The oligonucleotide sequences were listed in Table S1 (Additional file 1: Table S1). The miRNA overexpression vector pCMV-MIR519A (MI0003182) and the empty vector control were obtained from OriGene (Rockville, MD, USA). The miR-519a sponge and empty vector control were purchased from GeneChem (China). Transfections were performed using Lipofectamine 2000 reagent (cat. no. 11668-019; Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.