Investigating the role of BAG6 in protein quality control

Abstract

Cellular protein quality control is maintained by a network of chaperones, enzymes and adaptor molecules that mediate the ubiquitylation of target proteins, recognise their modified forms and promote their degradation. The BAG6 subunit, of the heterotrimeric BAG6 complex can recognise unfolded/hydrophobic polypeptides to act as both a "holdase" to prevent aggregation, a scaffold to recruit ubiquitylation machinery and a proteasomal delivery factor to facilitate substrate degradation. Studying the role of BAG6 during ER-associated degradation (ERAD) revealed that not only were normal levels of BAG6 necessary to promote efficient degradation of the model substrate opsin degron (OpD), but also for OpD ubiquitylation and the maintenance of global protein ubiquitylation. These data demonstrate that maintaining appropriate BAG6 levels is crucial for efficient cellular protein ubiquitylation and ERAD. Further endogenous BAG6 is required to maintain normal levels of CHIP and RNF126. Loss of BAG6, CHIP or RNF126 resulted in substrate-specific protein aggregation. Hence, a reduction in BAG6 and/or CHIP resulted in the aggregation of the ERAD substrate OpD, whilst a reduction in RNF126 led to the aggregation of the mislocalised protein (MLP), Op91. However, MLP aggregation following an RNF126 knockdown occurred only in the presence of BAG6. We conclude that BAG6 can act synergistically with CHIP to promote ERAD, and may function redundantly with an alternative pathway to mediate MLP degradation. To define additional cytosolic quality control factors, a proteomics screen was performed. In addition, to known regulators of protein quality control, including BAG6 and SGTA, we identified several type-II UBL domain proteins including members of the Ubiquilin family and RAD23A, together with the E3 ligases, MKRN1 and UBR4, components previously implicated in protein quality control processes. These preliminary findings suggest that multiple UBL-domain containing proteins and E3 ligases may contribute to the quality control of target substrates, providing a highly flexible and redundant system for protein clearance.

Date of Award	1 Aug 2015
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Simon Hubbard (Supervisor) & Stephen High (Supervisor)