The human BCa-derived cell lines 5637, RT112, UM-UC-5, UM-UC-9, VM-CUB-1, were obtained from DSMZ (Heidelberg, Germany), and UM-UC-3 from ATCC. UM-UC-3, UM-UC-5, UM-UC-9, VM-CUB-1 cells were cultured in DMEM (Life Technologies, Villebon-Sur-Yvette, France), whereas RT112 and 5637 cells were cultured in RPMI (Life Technologies). Media were supplemented with 10% fetal bovine serum (FBS, ThermoFisher Scientific, Courtaboeuf, France). Cells were kept at 37 °C, under an atmosphere containing 5% CO2. The identity of the cell lines used was checked by analyzing genomic alterations on comparative genomic hybridization arrays (CGH arrays) and sequencing genes known to be mutated: RAS, TP53, FGFR3 and PIK3CA. The cells were routinely checked for mycoplasma contamination.
Control and TYRO3-overexpressing cell lines were established by transfecting UM-UC-3 cells with a pCMV6-Entry vector (PS100001–Origene, Rockville, MD, USA) or a pCMV6-Entry vector encoding for the human TYRO3 open reading frame (RC208260 –Origene, Rockville, MD, USA) using lipofectamine 2000 (11668–027–ThermoFischer Scientific, Courtaboeuf, France) following the manufacturer’s protocol (3:1 DNA:lipofectamine 2000 ratio, 2 µg of DNA/well in 6-well plate). The selection of the transfected cell populations was performed using 1 mg/mL of Geneticin (10131035–ThermoFischer Scientific, Courtaboeuf, France).
Control and TYRO3-overexpressing cell lines were established by transfecting UM-UC-3 cells with a pCMV6-Entry vector (PS100001–Origene, Rockville, MD, USA) or a pCMV6-Entry vector encoding for the human TYRO3 open reading frame (RC208260 –Origene, Rockville, MD, USA) using lipofectamine 2000 (11668–027–ThermoFischer Scientific, Courtaboeuf, France) following the manufacturer’s protocol (3:1 DNA:lipofectamine 2000 ratio, 2 µg of DNA/well in 6-well plate). The selection of the transfected cell populations was performed using 1 mg/mL of Geneticin (10131035–ThermoFischer Scientific, Courtaboeuf, France).