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. 2012 Sep 4;122(10):3742–3754. doi: 10.1172/JCI62727

Figure 7. HSF1 knockdown impairs human MPNST cell growth and attenuates MAPK signaling.

Figure 7

(A) Differential efficacy of HSF1-targeting shRNA constructs. S462 cells were transduced with GSP or scrambled control or with hA9 or hA6 HSF1-targeting lentiviral supernatants and harvested for immunoblotting. (B) HSF1 knockdown impaired net growth and survival of MPNST cells. After plating, cells were transduced with viral supernatants as indicated. Relative viable cell number in each well was measured 4 days after viral transduction by resazurin dye reduction assay. Raw fluorescence data were normalized to values obtained in wells that underwent mock transduction (mean ± SD, n = 5). #P < 0.001, 2-way ANOVA. (C) Decreased levels of KSR1, p-ERK, HSP90α, and HSP72 after HSF1 knockdown. S462 cells were stably transduced with the indicated viral supernatants and harvested for immunoblotting. (D) Decreased levels of KSR1 and p-ERK after HSP90 inhibition. S462 cells were treated overnight with 1 μM geldanamycin (GA) and harvested for immunoblotting. (E) KSR1 knockdown induced apoptosis in MPNST cells. S462 cells were transfected with a nontargeting siRNA (NC) or 3 independent KSR1-targeting siRNAs at 25 nM final concentration. 2 days after transfection, cells were harvested for immunoblotting. (F) KSR1 knockdown impaired net growth and survival of MPNSTs. S462 cells plated in a 96-well format were transiently transfected without or with 25 nM siRNAs. Relative viable cell number in each well was measured as described in B 3 days after transfection (mean ± SD, n = 5). ***P < 0.001, 1-way ANOVA.