Structural and functional insights into the E3 ligase, RNF126

Abstract

RNF126 is an E3 ubiquitin ligase that collaborates with the BAG6 sortase complex to ubiquitinate hydrophobic substrates in the cytoplasm that are destined for proteasomal recycling. Composed of a trimeric complex of BAG6, TRC35 and UBL4A the BAG6 sortase is also associated with SGTA, a co-chaperone from which it can obtain hydrophobic substrates. Here we solve the solution structure of the RNF126 zinc finger domain in complex with the BAG6 UBL domain. We also characterise an interaction between RNF126 and UBL4A and analyse the competition between SGTA and RNF126 for the N-terminal BAG6 binding site. This work sheds light on the sorting mechanism of the BAG6 complex and its accessory proteins which, together, decide the fate of stray hydrophobic proteins in the aqueous cytoplasm.

Mice lacking WRB reveal differential biogenesis requirements of tail-anchored proteins in vivo

“…BAG6 is a component of the TA protein pre-targeting complex but is also known to be involved in the degradation of mislocalized proteins via the ubiquitin-proteasome system67. Together with our finding that Stx5 protein levels were lower in mammalian WRB knockout cells (Figs 1e,f and 2e,f) while those of Stx8 were nearly unchanged, this dual function of the BAG6-containing pre-targeting complex led us to test in HeLa cells whether Stx5 was targeted for proteasomal degradation in the absence of the TRC40 receptor.…”

“…A pre-targeting complex including the proteins BAG6, TRC35, and UBL4A receives newly synthesized TA proteins from the ribosome and transfers them to the cytosolic ATPase TRC405. BAG6 also actively promotes degradation of mislocalized proteins by targeting them to the ubiquitin-proteasome system67. TRC40 delivers TA proteins to a receptor formed by WRB (Tryptophan Rich Basic protein)8 and CAML (Calcium signal-modulating cyclophilin ligand)9, two integral membrane proteins localized at the ER.…”

Iruka Eliminates Dysfunctional Argonaute by Selective Ubiquitination of Its Empty State

“…However, unlike the manipulation of Iru levels, overexpression or depletion of CG7546, Get4, or CG7215 in S2 cells did not affect the ubiquitination of Ago1-YE ( Figures S2H and S2I). The Zinc domain of RNF126 and the Ubiquitin-like (Ubl) domain of Bag6 have been shown to interact in human cells (Krysztofinska et al, 2016;Rodrigo-Brenni et al, 2014), and deletion of the Zinc domain from Iru (Iru-DZinc) or of the Ubl domain from CG7546 (CG7546-DUbl) specifically abolished their interaction in S2 cells (Figures S2J and S2K). Nevertheless, Iru-DZinc, which cannot bind CG7546, was still able to bind (Figure S2L) and ubiquitinate ( Figure S2M) Ago1-YE similar to wildtype Iru.…”

SGTA interacts with the proteasomal ubiquitin receptor Rpn13 via a carboxylate clamp mechanism

“…SGTA and the BAG6 complex were initially discovered through their role in the biogenesis of tail-anchored membrane proteins and subsequently shown to regulate the ubiquitination and proteasomal degradation of MLPs23. The role of SGTA and BAG6 in both processes relies on multiple transient, and subtly discriminated, interactions with diverse binding partners and effectors45678910. These interactions contribute to a proposed BAG6/SGTA quality control cycle that can direct hydrophobic substrates towards either ubiquitination and proteasomal degradation (MLPs) or membrane insertion (tail-anchored proteins)2345111213.…”

“…It has a central TPR domain capable of direct interaction with both Hsp70 and Hsp90 chaperones, proteasomal subunits and a variety of disease-related proteins618, and a C-terminal domain capable of binding hydrophobic substrates19. Within this SGTA/BAG6 quality control cycle, BAG6 identifies hydrophobic MLPs and recruits an E3 ubiquitin ligase, RNF126, thereby facilitating their selective ubiquitination and entry into the pathway for proteasomal degradation813. Conversely, SGTA promotes substrate deubiquitination, thereby delaying the proteasomal degradation of MLPs45.…”