Knockdown of DHS mRNA in U87-MG and G55T2 cell lines by gene specific shRNAs.

<p>(A) Relative DHS-mRNA expression in G55T2 and U87-MG cells after lentiviral transduction with a scrambled shRNA (G55T2-ctrl and U87 ctrl) or DHS-specific shRNA pools normalized against GAPDH (mean±SD, <i>n = </i>3). (B) Immunoblot (30 µg total protein per sample) against DHS and β-Tubulin in G55T2 and U87-MG cells after lentiviral transduction with a scrambled shRNA (G55T2-ctrl and U87-ctrl) or DHS-specific shRNAs (G55T2 DHS-sh and U87 DHS-sh) after two days of puromycin selection. (C) Glioblastoma cell lines after transduction with a scrambled control shRNA or an shRNA-pool against DHS (100× magnification) with 100 µm scale bars. (D) Proliferation of transduced cells measured by trypan blue exclusion over five days. Significantly different cell numbers or DHS expression compared to controls are indicated (*:P<0.05; **:P<0.001). (E) Immunoblot results of U87-MG and G55T2 cells after transduction with a scrambled control shRNA (ctrl) or four different shRNAs against eIF-5A (30 µg total protein). Transduced cells were selected by incubation with puromycin for two days, cell lysates were obtained seven days post-selection. (F) Proliferative capacity of transduced cells with strong (A1-sh2 for U87-MG and A1-sh1 for G55T2) or moderate (A1-sh3 for U87-MG and A1-sh2 for G55T2) eIF-5A knockdown was determined by trypan blue exclusion cell counting over five days. Significantly different cell numbers compared to controls are marked by an asterisk (*:p<0.05).