Fig 2: Tumor compartment–specific expression of PSMB8 relates to clinicopathologic variables.(A) Representative β5i, MHC-I, and PA28β IHC analysis of normal breast ducts and lobules (i) and invasive breast tumors (ii, iii), illustrating cases that exemplify maintenance (ii) or selective loss (iii) of these class-I antigen presentation components. Cores are 1.0 mm i.d. and insets 140 μm2. (B) Contingency analysis of the relationships between β5i, MHC-I, and PA28β in triple-negative breast cancer (TNBC). Fisher’s exact test P-values indicated *P < 0.05; **P < 0.01; ***P < 0.001. (C) Contingency analysis of relationships between β5i, MHC-I, or PA28β, with TILs density (i), TILs PD-L1 positivity (ii), tumor cell PD-L1 positivity (iii), and the density of stromal APCs (sAPCs; iv). Fisher’s exact test P-values indicated *P < 0.05; **P < 0.01; ***P < 0.001. (D, E, F) Kaplan–Meier analysis of TNBCs stratified by β5i, MHC-I, or PA28β (i) or by TIL density after classifying TNBCs by their maintenance or loss of β5i, MHC-I, or PA28β (ii). HR, hazard ratio (95% confidence interval); log-rank P-values shown. (G) Change in β5i IHC scores in brain metastases (BrM) compared with matching primary breast cancers (BC). Overall numbers of cases exhibiting increases (↑), decreases (↓), or no change (–) are indicated. Paired, two-tailed t test P-value shown.

Fig 3: Oxyphylla A promotes PSMB8 expression and enhances proteasomal activity through the PKA/Akt/mTOR pathway. Inducible PC12/A53T-α-syn cells were treated with 1 μg/ml Dox for 24 h and then with Oxyphylla A (100 μM) for the indicated time periods. The expression ratios of phosphorylated PKA/total PKA (A), phosphorylated Akt/total Akt (B), phosphorylated p-mTOR/total mTOR (C) and phosphorylated p-p70s6k/total p70s6k (D) were detected by Western blotting analysis. PC12/A53T-α-syn cells were pretreated with or without PKA inhibitor H-89 (10 μM) for 1 h and then treated with Oxyphylla A (100 μM) for 30 min. The expression ratios of phosphorylated Akt/total Akt (E), phosphorylated p70S6K/total p70S6K (F) were analyzed by Western blotting. PC12/A53T-α-syn cells were pretreated with or without PKA inhibitor H-89 (10 μM) for 1 h and then treated with Oxyphylla A (100 μM) for another 24 h. The protein levels of PSMB8 were determined by Western blotting (G). Cellular chymotrypsin-like activity was determined by specific bioluminogenic enzyme substrates (H). Representative Western blotting and data analysis of three independent experiments are shown. Data are represented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 4: In vitro activation of the unfolded protein response (UPR) by bortezomib in relation to proteasome subunit expression.(A) Working model of UPR induction by three major signaling axes. BiP, binding immunoglobulin protein (GRP78). (B) qRT-PCR for ATF4 following bortezomib treatment. (B, C) Log2 fold-change (FC) in XBP1s, apoptosis markers, and NFκB at multiple time points after bortezomib treatment (B, C: qRT-PCR). (D) Inverse association between PSMB8/UPR induction and bortezomib sensitivity. (E) PSMB8 qRT-PCR (i) and bortezomib-induced cell death (ii) after pretreatment with IFN-γ. (F) IFN-γ–mediated induction of IRF1 and antigen processing genes in MDA-MB-468/MCF7 cells transfected with scrambled (SCR) or IRF1-specific siRNAs. P-values in this figure were from unpaired, two-tailed t tests: **P < 0.01; ***P < 0.001; ****P < 0.0001.

Fig 5: i-proteasome genes are induced in claudin-low TNBCs and coordinately expressed with unfolded protein response genes.(A, B) i:c Subunit expression ratios in breast cancer subtypes (METABRIC data, ANOVA test. *P < 0.05; ****P < 0.0001). (C) Gene set enrichment analysis plots showing running enrichment scores (ESs) skewed toward the PSMB8-correlated transcriptome. NES, normalized ES.