The Otubain YOD1 Is a Deubiquitinating Enzyme that Associates with p97 to Facilitate Protein Dislocation from the ER

Abstract

Summary YOD1 is a highly conserved deubiquitinating enzyme of the ovarian tumor (otubain) family, whose function has yet to be assigned in mammalian cells. YOD1 is a constituent of a multiprotein complex with p97 as its nucleus, suggesting a functional link to a pathway responsible for the dislocation of misfolded proteins from the endoplasmic reticulum. Expression of a YOD1 variant deprived of its deubiquitinating activity imposes a halt on the dislocation reaction, as judged by the stabilization of various dislocation substrates. Accordingly, we observe an increase in polyubiquitinated dislocation intermediates in association with p97 in the cytosol. This dominant negative effect is dependent on the UBX and Zinc finger domains, appended to the N- and C-terminus of the catalytic otubain core domain, respectively. The assignment of a p97-associated ubiquitin processing function to YOD1 adds to our understanding of p97’s role in the dislocation process.

Protein Folding and Quality Control in the ER

“…p97 binds to several ERAD components, including Derlin-1, VIMP (SelS), UBXD2 (Erasin), and UBXD8, and recruits several ubiquitin-chain modifiers including E3 ligases (gp78, HRD1, etc. ), chain elongation factors (Ufd2, E4 ubiquitin ligase), and deubiquitinases (YOD1, Ataxin-3, VCIP135) (Liang et al 2006;Mueller et al 2008;Schuberth and Buchberger 2008;Ernst et al 2009). Ubiquitinated substrates are transferred to the proteasome by shuttle proteins, known as HR23A/B or Ubiquilin-1 (Rad23 and Dsk2 in yeast), which contain ubiquitinassociated (UBA) and ubiquitin-like (UBL) domains that bind to polyubiquitin chains and the proteasome subunits (Rpn10/13, Rpt5), respectively (Deveraux et al 1994;Lam et al 2002;Raasi and Wolf 2007;Husnjak et al 2008;Finley 2009;Lim et al 2009).…”

Control of p97 function by cofactor binding

“…So far, 13 UBX domain-containing proteins have been identified in mammals, all of which, with the exception of UBXD1 (see below), have been demonstrated to bind to p97 [46,47,49]. In addition, the related UBXL domain has been identified in a few p97 cofactors including the NPL4 subunit of the heterodimeric UFD1-NPL4 cofactor [50] and the two DUBs OTU1/YOD1 [51] and VCIP135 [52]. Due to their higher resolution, we will focus the following discussion on the UBX and UBXL domains of FAF1 and OTU1, respectively.…”

A Viral Deubiquitylating Enzyme Restores Dislocation of Substrates from the Endoplasmic Reticulum (ER) in Semi-intact Cells

“…The ubiquitin-specific protease Yod1 plays a role in clearing the ER of several misfolded substrates (13,14). Expression of a dominant negative version of Yod1 (the YodC160S mutant, which lacks catalytic activity) leads to accumulation in the ER of substrates (in their nonubiquitylated state) that would otherwise have been discharged and destroyed (13).…”

“…Expression of a dominant negative version of Yod1 (the YodC160S mutant, which lacks catalytic activity) leads to accumulation in the ER of substrates (in their nonubiquitylated state) that would otherwise have been discharged and destroyed (13). We hypothesized that a failure to remove ubiquitin from a dislocation substrate en route from the dislocon to the next station in the dislocation pathway would stall the substrate and block all further dislocation.…”