Fig. 4 |. Genetic screens reveal increased ER stress and decreased Notch signaling following ZIP7 siRNA knockdown.

a, Gene on-target activity in the ERSE-Luc assay agonist side (increased signaling) was plotted as robust z-score (activity of the 75th percentile siRNA per gene) as a function of RSA (significance of enrichment) for NVS-ZP7–4 over DMSO control. Color was set to depict the number of siRNAs per gene with significant activity (robust z-score > + 2). Dotted lines represent stringent (green) and loose (red) thresholds of significance of enrichment, determined on the basis of randomized dataset analysis. b, The impact of the individual ZIP7 siRNAs (log₂ fold change; luminescence for test siRNA over median luminescence for all siRNAs) in the ERSE-Luc assay is shown for each of the three screening conditions (DMSO, 20 nM NVS-ZP7–4, 20 nM NVS-ZP7–4 over DMSO). c, Quantitation of ZIP7 mRNA expression after treatment of HSC-3 cells with control (pGL2) or four independent ZIP7 siRNAs for 24 h. Data are representative of eight technical replicates from one biological sample represented in a box-plot graph. d, NVS-ZP7–4 dose response in ERSE-Luc assay in combination with siRNA knockdown with two independent ZIP7 siRNAs or a control siRNA (pGL2). Error bars represent s.d. of the mean from six biological replicates (n = 6) in an individual experiment. Each experiment was performed three independent times. e, NVS-ZP7–4 dose response in Notch signaling HES-luciferase assay in combination with siRNA knockdown with two independent ZIP7 siRNAs or a control siRNA (pGL2). Error bars represent the s.d. of the mean from six biological replicates (n = 6) in an individual experiment. Each experiment was performed three independent times.