Supplemental Information Recruitment and Activation of a Lipid Kinase by Hepatitis C Virus NS 5 A Is Essential for Integrity of the Membranous Replication Compartment

Cell lines, viruses and plasmid constructs Subgenomic replicon cell lines JFH-1 and Con1ET expressing the FLuc gene have been described recently (Jo et al., 2009; Lohmann et al., 2003; Vrolijk et al., 2003). Immunofluorescence studies were performed using cells carrying a selectable JFH-1 subgenomic replicon referred to as neo-sgJFH-1 (Schaller et al., 2007). Huh7-Lunet cells served as mock control (Friebe et al., 2005). Transfections of T7 promotor driven constructs were performed in Huh7-Lunet cells stably expressing the T7 polymerase (Huh7-Lunet/T7) (Appel et al., 2005). Cells with stable knock-down of PI4KIIIα (sh-PI4KIIIα) or non-targeting control cells (sh-NT) were prepared according to a recently published protocol (Kaul et al., 2009). The shRNA targeting sequences were 5‟-AGG TAC GAC AAG ATG GGC TAT-3‟ (PI4KIIIα) and 5‟-TCT CGC TTG GGC GAG AGT AAG-3‟ (NT).