Article Title: CRISPR-Cas9 Screening of Kaposi’s Sarcoma-Associated Herpesvirus-Transformed Cells Identifies XPO1 as a Vulnerable Target of Cancer Cells
Figure Lengend Snippet: Identification of XPO1 as a vulnerable gene of KSHV-transformed KMM cells and of cell lines of gastric cancer AGS and liver cancer HUH7 cells. (A and B) XPO1 expression in MM cells and in MM cells infected by KSHV (KMM) and by mutant viruses with a deletion of vFLIP (ΔvFLIP), vCyclin (ΔvCyclin), or a cluster of 10 pre-miRNAs (ΔmiR) analyzed by reverse transcription-quantitative PCR (qRT-PCR) (A) and Western blotting (B). (C) Expression of XPO1 analyzed by qRT-PCR in MM cells, KMM cells, ΔmiR cells, or ΔmiR cells complemented with individual KSHV pre-miRNAs (K1 to K12). (D) Expression of XPO1 following siRNA knockdown analyzed by Western blotting in MM cells and KMM cells. (E) Expression of XPO1 following siRNA knockdown analyzed by Western blotting in AGS and HUH7 cells. (F) Analysis of cell proliferation following siRNA knockdown of XPO1 in MM, KMM, AGS, and HUH7 cell lines. (G) Analysis of cell proliferation following treatment with DMSO or KPT-8602 at different concentrations for 3 days in MM, KMM, AGS, and HUH7 cells. (H) Formation of colonies in soft agar following treatment with DMSO or KPT-8602 at 1 µM in KMM, AGS, and HUH7 cells. Representative fields are shown, and efficiencies of colony formation are presented.
Article Snippet: Primary antibodies to β-actin (Santa Cruz), p62 (CST), SIRT1 (CST), Cas9-hemagglutinin (Cas9-HA; Santa Cruz), PML (CST), XPO1 (CST), p53 (CST), and phospho-p53 (CST) were used.
Techniques: Transformation Assay, Expressing, Infection, Mutagenesis, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot