Curcumin inhibits proteasome activity in triple-negative breast cancer cells through regulating p300/miR-142-3p/PSMB5 axis

Background: Curcumin functions as a proteasome inhibitor. However, the molecular mechanisms behind this action need more detailed explanations. Purpose: This study aimed to investigate the inhibitory effect of curcumin on 20S proteasome activity and to elucidate its exact mechanism in triple-negative breast cancer (TNBC) MDA-MB-231 cells. Methods: Proteasomal peptidase activities were assayed using synthetic fluorogenic peptide substrates. Knockdown or overexpression of microRNA (miRNA or miR) or protein was used to investigate its functional effect on downstream cellular processes. BrdU (5-bromo-2'-deoxyuridine) assay was performed to identify cell proliferation. Western blot and quantitative real-time PCR(qRT-PCR) were carried out to determine protein abundance and miRNA expression, respectively. Correlations between protein expressions, miRNA levels, and proteasome activities were analyzed in TNBC tissues. Xenograft tumor model was performed to observe the in vivo effect of curcumin on 20S proteasome activity. Results: Curcumin significantly reduced PSMB5 protein levels, accompanied with a reduction in the chymotrypsin-like (CT-l) activity of proteasome 20S core. Loss of PSMB5 markedly inhibited the CT-1 activity of 20S proteasome. Furthermore, curcumin treatment significantly elevated miR-142-3p expression. PSMB5 was a direct target of miR-142-3p and its protein levels were negatively regulated by miR-142-3p. Moreover, histone acetyltransferase p300 suppressed miR-142-3p expression. Overexpression of p300 mitigated the promotive effect of curcumin on miR-142-3p expression. The correlations among p300 abundances, miR-142-3p levels, PSMB5 expressions, and the CT-1 activities of 20S proteasome were evidenced in TNBC tissues. In addition, loss of p300 and PSMB5 reduced cell proliferation. Inhibition of miR-142-3p significantly attenuated the inhibitory impact of curcumin on cell proliferation. These curcumin-induced changes on p300, miR-142-3p, PSMB5, and 20S proteasome activity were further confirmed in in vivo solid tumor model. Conclusion: These findings demonstrated that curcumin suppressed p300/miR-142-3p/PSMB5 axis leading to the inhibition of the CT-1 activity of 20S proteasome. These results provide a novel and alternative explanation for the inhibitory effect of curcumin on proteasome activity and also raised potential therapeutic targets for TNBC treatment.