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. 2021 Oct 7;220(12):e202103087. doi: 10.1083/jcb.202103087

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© 2021 Herbst et al.

This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).

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Figure 3. — PDCD4 undergoes proteasome-mediated degradation, not nuclear export, following neuronal stimulation. (A) Top: Schematic of CRM1-mediated nuclear export inhibitor LMB experiments. Bottom: Violin plots of normalized nuclear PDCD4 ICC intensity. Basal, n = 109; TTX, n = 109; Bic, n = 109; LMB-basal, n = 106; LMB-TTX, n = 100; and LMB-Bic, n = 86 cells from 3 sets of cultures. Basal median = 1.00; TTX median = 1.08; Bic median = 0.59; LMB-basal median = 1.30; LMB-TTX median = 1.16; LMB-Bic median = 0.52. Basal versus Bic, P < 0.0001; TTX versus Bic, P < 0.0001; LMB-basal versus LMB-Bic, P < 0.0001; LMB-TTX versus LMB-Bic, P < 0.0001. (B) Top: Schematic of proteasome inhibitor Epox experiments. Bottom: Violin plots of normalized nuclear PDCD4 ICC intensity. Basal, n = 113; TTX, n = 106; Bic, n = 103; Epox-basal, n = 107; Epox-TTX, n = 94; and Epox-Bic, n = 86 cells from 3 sets of cultures. Basal median = 1.00; TTX median = 1.18; Bic median = 0.62; Epox-basal median = 0.97; Epox-TTX median = 0.99; Epox-Bic median = 0.88. Basal versus Bic, P < 0.0001; TTX versus Bic, P < 0.0001; Epox-basal versus Epox-Bic, P = 0.1662; Epox-TTX versus Epox-Bic, P = 0.0174. (C) Top: Schematic of proteasome inhibitor Bort experiments. Bottom: Violin plots of normalized nuclear PDCD4 ICC intensity. Basal, n = 131; TTX, n = 111; Bic, n = 120; Bort-basal, n = 100; Bort-TTX, n = 98; and Bort-Bic, n = 116 cells from three sets of cultures. Basal median = 1.00; TTX median = 1.09; Bic median = 0.60; Bort-basal median = 1.00; Bort-TTX median = 1.04; Bort-Bic median = 0.94. Basal versus Bic, P < 0.0001; TTX versus Bic, P < 0.0001; Bort-basal versus Bort-Bic, P = 0.6224; Bort-TTX versus Bort-Bic, P = 0.2648. (D) Top: Schematic of MLN experiments. MLN inhibits neddylation, preventing the activation of Cullin-RING E3 ubiquitin ligases. Bottom: Violin plots of normalized nuclear PDCD4 ICC intensity. Basal, n = 130; TTX, n = 120; Bic, n = 120; MLN-basal, n = 115; MLN-TTX, n = 108; and MLN-Bic, n = 97 cells from three sets of cultures. Basal median = 1.00; TTX median = 1.11; Bic median = 0.64; MLN-basal median = 1.05; MLN-TTX median = 0.93; MLN-Bic median = 0.95. Basal versus Bic, P < 0.0001; TTX versus Bic, P < 0.0001; MLN-basal versus MLN-Bic, P = 0.329; MLN-TTX versus MLN-Bic, P = 1. *, P < 0.05; ****, P < 0.0001; Mann-Whitney U test with Bonferroni correction.