Constitutive Activation of p62/Sequestosome-1-Mediated Proteaphagy Regulates Proteolysis and Impairs Cell Death in Bortezomib-Resistant Mantle Cell Lymphoma

SIMPLE SUMMARY: To decipher the molecular mechanism underlying the resistance of a significant fraction of mantle cell lymphoma (MCL) patients to the first-in-class proteasome inhibitor bortezomib (BTZ), we have characterized the ubiquitin-related proteome (i.e., ubiquitome) of a set of MCL cell lines with different degrees of sensitivity to the drug by coupling a tandem ubiquitin-binding entity (TUBE) approach to mass spectrometry, followed by phenotypic and functional validations in both in vitro and in vivo models of MCL. We identified an enrichment of autophagy–lysosome system (ALS) components in BTZ-resistant cells, which was associated with constitutive intracellular inactivation of proteasome subunits by a process called proteaphagy. Blockade of this phenomenon by the pharmacological or genetic inactivation of the autophagy receptor p62/SQSTM1 reactivated normal proteasomal activity and restored the BTZ antitumor effect in in vitro and in vivo models of BTZ resistance. ABSTRACT: Protein ubiquitylation coordinates crucial cellular events in physiological and pathological conditions. A comparative analysis of the ubiquitin proteome from bortezomib (BTZ)-sensitive and BTZ-resistant mantle cell lymphoma (MCL) revealed an enrichment of the autophagy–lysosome system (ALS) in BTZ-resistant cells. Pharmacological inhibition of autophagy at the level of lysosome-fusion revealed a constitutive activation of proteaphagy and accumulation of proteasome subunits within autophagosomes in different MCL cell lines with acquired or natural resistance to BTZ. Inhibition of the autophagy receptor p62/SQSTM1 upon verteporfin (VTP) treatment disrupted proteaphagosome assembly, reduced co-localization of proteasome subunits with autophagy markers and negatively impacted proteasome activity. Finally, the silencing or pharmacological inhibition of p62 restored the apoptosis threshold at physiological levels in BTZ-resistant cells both in vitro and in vivo. In total, these results demonstrate for the first time a proteolytic switch from the ubiquitin–proteasome system (UPS) to ALS in B-cell lymphoma refractory to proteasome inhibition, pointing out a crucial role for proteaphagy in this phenomenon and paving the way for the design of alternative therapeutic venues in treatment-resistant tumors.