Figure 1 | Cell Death & Disease

Figure 1

From: Functional genetics-directed identification of novel pharmacological inhibitors of FAS- and TNF-dependent apoptosis that protect mice from acute liver failure

Figure 1

RNAi-based phenotypic screening approach used to identify modulators of FAS- and TNF-induced apoptosis. (a) Pathway Decipher genes (n=5046) separated into annotated functional groups, with the sizes of different pie segments indicating the relative frequency of genes within the corresponding groups. (b) Lentiviral expression vector used for construction of the 27 K pooled shRNA Library targeting Pathway Decipher genes. (c) Construction of the 27 K Pathway Decipher shRNA Library and scheme of shRNA library screening for inhibition of FASL- or TNF-induced apoptosis. (d) Canonical FAS and TNF receptor-mediated apoptotic pathways. (e) Effects of shRNAs targeting genes for canonical FAS and TNF pathway members on viability of cells treated with FASL or TNF. shRNAs (two per target gene) were cloned into the lentiviral expression vector shown in b in an arrayed library format and transduced into individual HeLa cell populations. After puromycin selection, transduced cells were exposed to FASL (50 ng/ml, dark gray bars) or TNF (50 ng/ml, light gray bars) in the presence of cycloheximide (2.5 μg/ml). Cell viability was measured 24 h later by Resazurin assay, with fluorescence units quantitatively correlated with cell viability. shRNAs against luciferase (LUC) were used as negative controls. The bars indicate relative survival of cells (%) versus untreated control cells

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