title: A plastic SQSTM1/p62-dependent autophagic reserve maintains proteostasis and determines proteasome inhibitor susceptibility in multiple myeloma cells.
abstract: Multiple myeloma (MM) is the paradigmatic proteasome inhibitor (PI) responsive cancer, but many patients fail to respond. An attractive target to enhance sensitivity is (macro)autophagy, recently found essential to bone marrow plasma cells, the normal counterpart of MM. Here, integrating proteomics with hypothesis-driven strategies, we identified the autophagic cargo receptor and adapter protein, SQSTM1/p62 as an essential component of an autophagic reserve that not only synergizes with the proteasome to maintain proteostasis, but also mediates a plastic adaptive response to PIs, and faithfully reports on inherent PI sensitivity. Lentiviral engineering revealed that SQSTM1 is essential for MM cell survival and affords specific PI protection. Under basal conditions, SQSTM1-dependent autophagy alleviates the degradative burden on the proteasome by constitutively disposing of substantial amounts of ubiquitinated proteins. Indeed, its inhibition or stimulation greatly sensitized to, or protected from, PI-induced protein aggregation and cell death. Moreover, under proteasome stress, myeloma cells selectively enhanced SQSTM1 de novo expression and reset its vast endogenous interactome, diverting SQSTM1 from signaling partners to maximize its association with ubiquitinated proteins. Saturation of such autophagic reserve, as indicated by intracellular accumulation of undigested SQSTM1-positive aggregates, specifically discriminated patient-derived myelomas inherently susceptible to PIs from primarily resistant ones. These aggregates correlated with accumulation of the endoplasmic reticulum, which comparative proteomics identified as the main cell compartment targeted by autophagy in MM. Altogether, the data integrate autophagy into our previously established proteasome load-versus-capacity model, and reveal SQSTM1 aggregation as a faithful marker of defective proteostasis, defining a novel prognostic and therapeutic framework for MM.
authors.affiliation: a San Raffaele Scientific Institute; Division of Genetics and Cell Biology ; Milan , Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Milan
authors.forename: Tommaso
authors.initials: T
authors.lastname: Perini
authors.forename: Massimo
authors.initials: M
authors.lastname: Resnati
authors.forename: Ugo
authors.initials: U
authors.lastname: Orfanelli
authors.forename: Laura
authors.initials: L
authors.lastname: Oliva
authors.forename: Andrea
authors.initials: A
authors.lastname: Raimondi
authors.forename: Paolo
authors.initials: P
authors.lastname: Cascio
authors.forename: Angela
authors.initials: A
authors.lastname: Bachi
authors.forename: Magda
authors.initials: M
authors.lastname: Marcatti
authors.forename: Fabio
authors.initials: F
authors.lastname: Ciceri
authors.forename: Simone
authors.initials: S
authors.lastname: Cenci
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D001343:Autophagy / Q000187:drug effects*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D019610:Cytoprotection / Q000187:drug effects; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure; D006706:Homeostasis / Q000187:drug effects*; D006801:Humans; D009101:Multiple Myeloma / Q000378:metabolism* / Q000473:pathology*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors / Q000494:pharmacology*; D066329:Protein Aggregates / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D059748:Proteolysis / Q000187:drug effects; D000071456:Sequestosome-1 Protein; D057149:Ubiquitinated Proteins / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D061988:Proteasome Inhibitors; D066329:Protein Aggregates; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D057149:Ubiquitinated Proteins; D046988:Proteasome Endopeptidase Complex
keywords: APC, allophycocyanin; Ab, antibody; BM, bone marrow; Baf A1, bafilomycin A1; Btz, bortezomib; ER, endoplasmic reticulum; ERGIC, ER-Golgi intermediate compartment; GO, gene ontology; HCQ, hydroxychloroquine; IP, immunoprecipitation; Ig, immunoglobulin; LC-MS/MS, liquid chromatography–tandem mass spectrometry; MM, multiple myeloma; PBS, phosphate-buffered saline; PC, plasma cell; PI, proteasome inhibitor; Rapa, rapamycin; SILAC, stable isotope labeling in cell culture; SQSTM1; UPR, unfolded protein response; UPS, ubiquitin-proteasome system; Ub, ubiquitin; aggregate; amc, 7-amino-4-methylcoumarin; autophagy; bortezomib; endoplasmic reticulum; multiple myeloma; p62; pAb, polyclonal antibody; plasma cells; proteasome; proteasome inhibitors; proteostasis; ubiquitin