Interactome screening implicates BAG6 as a suppressor of UBQLN2 misfolding in ALS/FTD.

Ubiquilin-2 (UBQLN2) is a ubiquitin (Ub)-binding shuttle protein that is mutated in X-linked forms of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). ALS/FTD-linked mutations in UBQLN2 disrupt its conformation, increasing its tendency to form cytoplasmic aggregates that may disrupt cellular regulation through loss-of-function (LOF) and gain-of-function (GOF) effects. Here, we performed quantitative mass spectrometry (MS)-based interactome analysis of wild-type (UBQLN2WT) and ALS-mutant UBQLN2 (UBQLN2ALS) proteins using inducible pluripotent stem cells (iPSCs) and induced motor neurons (iMNs). Proteins showing enhanced association with UBQLN2ALS proteins included PEG10, a known degradation target of UBQLN2, and BAG6, a chaperone involved in the triage of mislocalized proteins (MLPs). BAG6 knockdown inhibited the solubility recovery of both UBQLN2WT and UBQLN2ALS proteins following heat stress (HS), suggesting it functions as a UBQLN2 holdase. In addition, knockdown of BAG6 or knockout of UBQLN2 led to PEG10 accumulation, implicating both in PEG10 turnover; however, neither BAG6 nor UBQLN2 was required for PEG10 degradation in response to HS. A highly aggregation-prone UBQLN24XALS mutant harboring four different ALS-associated mutations showed increased PEG10 binding and modestly delayed PEG10 turnover while PEG10 degradation was not significantly different between UBQLN2WT and iPSCs expressing a UBQLN2P497H clinical mutant. The combined findings implicate BAG6 as a UBQLN2 holdase and identify a suite of proteins whose altered binding may contribute to pathologic changes in UBQLN2-associated ALS/FTD.