Mda mb231 mda 231

The human breast cancer cell lines MDA-MB-231 (MDA231) and MCF7 were purchased from American Type Culture Collection (ATCC). Genetically modified breast cancer cell lines (MDA.Gluc and MCF7.Gluc) were generated by transfection with Gaussian-luc cDNA construct to promote the constitutive expression of Gaussia luciferase in neomycin resistant cells. The stable Rab27a knockdown cell line (MDA.Rab27a.shRNA) was established by Rab27a shRNA lentivirus (sc-41834-v, Santa Cruz Biotechnology, Bio-Strategy Pty Limited, Australia) transduction using Polybrene® (sc-134220, Santa Cruz Biotechnology, Bio-Strategy Pty Limited, Australia) in puromycin resistant cells. Transient Rab27a knockdown in MDA231 cells was established by Rab27a siRNA (sc-41834, Santa Cruz Biotechnology, Bio-Strategy Pty Limited, Australia) transfection using Lipofectamine™ RNAiMAX Transfection Reagent (Invitrogen, Thermo Fisher Scientific, Australia PTY LTD). Successful knockdown was confirmed by western blot. All cell lines were cultured in Dulbecco’s modified Eagle’s medium (DMEM) (1965118, Gibco, Thermo Fisher Scientific, Australia PTY LTD) supplemented with 10% fetal calf serum (FCS) (HyCloneTM, GE Healthcare Life Sciences, USA), 10 µg/mL penicillin and 100 µg/mL streptomycin (Invitrogen, Thermo Fisher Scientific, Australia PTY LTD). Cell cultures were grown at 37 °C with 10% CO₂ in a humidified atmosphere.

Human breast cancer cell lines MDA-MB-231 (MDA-231), BT549 and human embryonic kidney 293T (HEK-293T) were purchased from the American Type Culture Collection (ATCC, Manassas, VA, United States). MDA-231 and BT549 cells were grown in RPMI-1640 medium (Hyclone, South Logan, UT, United States); HEK-293T cells were cultured in DMEM medium, and all media were supplemented with 10% fetal bovine serum (HyClone) under standard culture conditions (5% CO₂ at 37 °C).

The human embryonic kidney 293 T cell line, human breast cancer cell lines MDA-MB-231 (MDA231), T47D and MCF7 were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA). IMP1 knockdown T47D cells, MDA231/IMP1-GFP and MDA231/GFP cell lines were previously generated [36 (link), 37 (link)]. UCA1 overexpression and knockdown MDA231 and T47D stable cell lines were established by stable infection of lentivirus expressing UCA1 or UCA1-short hairpin RNA (shRNA). Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) (Thermo Fisher Scientific, USA) supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin, and 100 μg/ml streptomycin at 37 °C in a humid environment with 5% CO₂.