Cdc48pNpl4p/Ufd1p Binds and Segregates Membrane-Anchored/Tethered Complexes via a Polyubiquitin Signal Present on the Anchors

Abstract

Cdc48p is an abundant and conserved member of the AAA ATPase family of molecular chaperones. Cdc48p performs ubiquitin-selective functions, which are mediated by numerous ubiquitin binding adaptors, including the Npl4p-Ufd1p complex. Previous studies suggest that Cdc48p-containing complexes carry out many biochemical activities, including ubiquitination, deubiquitination, protein complex segregation, and targeting of ubiquitinated substrates to the proteasome. The molecular mechanisms by which Cdc48p-containing complexes participate in these processes remain poorly defined. We show here by using physiologically relevant Cdc48p substrates (i.e., endoplasmic membrane-associated/tethered dimers of Mga2p and Spt23p) and in vitro systems with purified proteins that Cdc48p(Npl4p/Ufd1p) binds to and promotes segregation of the tethered proteins via a polyubiquitin signal present on the membrane-bound proteins. Mobilization does not involve retrotranslocation of the associated anchors. These results provide biochemical evidence that Cdc48p(Npl4p/Ufd1p) functions as a polyubiquitin-selective segregase and that a polyubiquitin-Cdc48p pathway modulates protein interactions at cell membranes.

A dual signalling pathway for the hypoxic expression of lipid genes, dependent on the glucose sensor Rag4, is revealed by the analysis of the KlMGA2 gene in Kluyveromyces lactis

“…Western blot analysis of crude extracts from the K. lactis-transformed clones (Fig. 6a) showed a maturation pattern of Mga2 similar to that reported in S. cerevisiae (Shcherbik & Haines, 2007), involving ubiquitination and proteasomal proteolysis.…”

The Ubiquitin–Proteasome System of Saccharomyces cerevisiae

“…Cdc48-Ufd1-Npl4 binds and mobilizes the ubiquitylated and processed p90, separating it from the unprocessed Spt23. These data suggested that Cdc48 acts as a segregase to disassemble protein complexes (Rape et al 2001;Jentsch and Rumpf 2007;Shcherbik and Haines 2007).…”

AAA ATPase p97/VCP is essential for TRIM21-mediated virus neutralization

“…Most recently, it has been shown that VCP is recruited to stalled proteasomes and relieves their (experimentally induced) impairment (21). It has been suggested that ATP hydrolysisdriven extraction of proteins from larger complexes or membranes, as well as unfolding of proteins by VCP upon recruitment to conjugated (poly)ubiquitin may be the common mechanism underlying the diverse cellular roles of VCP (13,16,22).The energy-dependent, ubiquitin-selective segregase and unfoldase activity of VCP prompted us to investigate whether it has a role in ADIN. The results presented here demonstrate that VCP is an essential cofactor for ADIN and that the ATPase activity of VCP is required when ADIN mediates proteasomal degradation of a challenging substrate such as an adenovirus particle.…”

“…Most recently, it has been shown that VCP is recruited to stalled proteasomes and relieves their (experimentally induced) impairment (21). It has been suggested that ATP hydrolysisdriven extraction of proteins from larger complexes or membranes, as well as unfolding of proteins by VCP upon recruitment to conjugated (poly)ubiquitin may be the common mechanism underlying the diverse cellular roles of VCP (13,16,22).…”