Colon Cancer Cell Lines and Knockdowns

Human colon cancer cell lines DLD1 and HCT116 cells were cultured in DMEM and McCoy’s 5A supplemented with 10% fetal bovine serum (FBS, Sigma-Aldrich, MO, USA) and 1% penicillin–streptomycin, respectively. These cells were purchased from ATCC and authenticated using short tandem repeat (STR) DNA profiling in March 2016 (Genetica, OH, USA). Primary colon cancer Pt130 cells were established from patient-derived xenografts (PDX) as described previously^{40 (link)}. Briefly, this cell line, obtained from a 63-year old male with moderately differentiated colonic adenocarcinoma, contains BRAF (V600E) and TP53 mutations and was authenticated using short tandem repeat (STR) DNA profiling as a unique cancer cell line (Genetica). Stable SREBP1, SREBP2 and SCAP knockdown cells were generated using lentivirus-based RNAi^{40 (link)–42 (link)}. The shRNA targeting sequences for SREBP1 (also named SREBF1) are as the following: 5′-GCCATCGACTACATTCGCTTT-3′ (a) and 5′-CCAGAAACTCAAGCAGGAGAA-3′ (b); for SREBP2 (also named SREBF2): 5′-CCTCAGATCATCAAGACAGAT-3′ (a) and 5′-GACCTGAAGATCGAGGACTTT-3′ (b); and for SCAP: 5′-GCTCAACGGTTCCCTTGATTT-3′ (a) and 5′-GCTCTGGTGTTCTTGGACAAA-3′ (b). The non-targeting control shRNA lentivirus plasmid (MISSION, SHC002) was obtained from Sigma-Aldrich (MO, USA). Fatostatin and 25-HC were purchased from Sigma-Aldrich.