In black flat-bottom 96-well plates (BD Biosciences), cells were plated (.10,000 cells/well) and infected as described in the methods below. Following infection, cells were fixed in 4% formaldehyde v/v diluted in PBS and blocked in PBS containing 3% BSA, 0.3% Triton X-100, and 10% FBS. Cells were first probed with HCV anti-core antibody (1:500) in PBS containing 3% BSA and 0.3% Triton X-100 (Binding Buffer), then incubated with Alexa Fluor-568-conjugated donkey anti-mouse secondary antibody (1:1000) and 10 mg/ml Hoechst dye. Cells were analyzed by a quantitative, high-throughput, fluorescence microscope system called the Cellomics ArrayScan VTI High Content Screening (HCS) Reader (Thermo Scientific) using the software Target Activation BioApplication (TABA). TABA was used to count the total number of cells (Hoechst-stained nuclei) and the percentage of those cells that were expressing HCV core (positive signal at 568 nm wavelength). . Nuclear fractionation of cell lysates was performed at 4uC as described previously with modifications [64, 66] . Cells were harvested in 400 ml buffer C (10 mM HEPES/KOH, pH 7.6, 10 mM KCl, 1.5 mM MgCl 2 , 1 mM EDTA, 1 mM EGTA, 250 mM sucrose) containing 1 X complete protease inhibitor cocktail (Roche). To shear DNA, the cells were passed 20 times through a 23-gauge needle. The lysate was centrifuged at 1100 x g for 7 minutes and the resulting supernatant was centrifuged again at 25000 x g for 60 minutes to obtain the membrane pellet. The pellet containing membranebound SREBP-1 was re-suspended in 75 ml of SDS-lysis buffer (10 mM Tris HCl, 100 mM NaCl, 1% SDS, 1 mM EDTA, 1 mM EGTA, pH 6.8). The pellet from the 1100 x g spin was resuspended in 100 ml buffer D (20 mM HEPES/KOH, 420 mM NaCl, 1.5 mM MgCl 2 , 2.5% glycerol, 1 mM EDTA, 1 mM EGTA, pH 7.6) containing 1 X complete protease inhibitor cocktail. Nuclear pellets were rocked for 1 hour, after which the samples were centrifuged at 25000 x g for 60 minutes to obtain the clarified supernatant containing the nuclear fraction.