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. 2021 Feb 20;19(2):117. doi: 10.3390/md19020117

Figure 5.

Figure 5

PNSA inhibits the protein expressions and activations in the signaling pathway by targeting heat shock protein 90 (HSP90). (A) Knockdown of HSP90 causes changes in protein expressions and activations. MDA-MB-231 cell line transfected with Si-RNA or Si-NC were cultured in 6-well plates for 48 h. Protein expressions and activations were assessed by Western blotting. (B) PNSA degrades HSP90 clients EGFR and C-Myc via proteasome pathway. MDA-MB-231 cells were treated with PNSA or/and MG132 (protease inhibitor) for 12 h. Protein levels were assessed by Western blotting. GAPDH was used as a loading control. (C) Effect of PNSA on HSP90 protein stabilization. MDA-MB-231 cells were treated with 2 μM PNSA for 3 h before heated at different temperatures. Western blotting was used to determine the protein levels (top panel). The protein band density was quantified by normalization to β-Tubulin (bottom panel). (D) Effects of PNSA and 17-AAG on the expressions of HSP70 and HSP90. MDA-MB-231 cells were treated with PNSA (0.5, 1, 2 μM) or 4 μM 17-AAG for 12 h. Proteins levels were analyzed by Western blotting. GAPDH was used as a loading control. The bar graph represents the average ± SD of at least three independent experiments. * p < 0.05; ** p < 0.01; ns, not significant (relative to DMSO-treated cells).