ESCRT ubiquitin-binding domains function cooperatively during MVB cargo sorting

Shields, S. Brookhart; Oestreich, Andrea J.; Winistorfer, Stanley C.; Nguyen, Doris; Payne, Johanna A.; Katzmann, David J.; Piper, Robert C.

doi:10.1083/jcb.200811130

Cited by 100 publications

(123 citation statements)

References 38 publications

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“…In contrast, additional UBDs may be present in other metazoan ESCRT complexes. Based on recent evidence that yeast ESCRT-I harbors an addition UBD in its Mvb12p subunit (30), it is possible that additional UBDs await characterization in other ESCRT complexes. Analysis of the interactions between intact ESCRT-I or -II and ubiquitin will be necessary to ultimately resolve this issue.…”

Section: Discussionmentioning

confidence: 99%

ESCRT-0 Assembles as a Heterotetrameric Complex on Membranes and Binds Multiple Ubiquitinylated Cargoes Simultaneously

Mayers¹,

Fyfe

Schuh³

et al. 2011

Journal of Biological Chemistry

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The ESCRT machinery consists of multiple protein complexes that collectively participate in the biogenesis of multivesicular endosomes (MVEs). The ESCRT-0 complex is composed of two subunits, Hrs and STAM, both of which can engage ubiquitinylated substrates destined for lysosomal degradation. Here, we conduct a comprehensive analysis of ESCRT-0:ubiquitin interactions using isothermal titration calorimetry and define the affinity of each ubiquitin-binding domain (UBD) within the intact ESCRT-0 complex. Our data demonstrate that ubiquitin binding is non-cooperative between the ESCRT-0 UBDs. Additionally, our findings show that the affinity of the Hrs double ubiquitin interacting motif (DUIM) for ubiquitin is more than 2-fold greater than that of UBDs found in STAM, suggesting that Hrs functions as the major ubiquitin-binding protein in ESCRT-0. In vivo, Hrs and STAM localize to endosomal membranes. To study recombinant ESCRT-0 assembly on lipid bilayers, we used atomic force microscopy. Our data show that ESCRT-0 forms mostly heterodimers and heterotetramers of Hrs and STAM when analyzed in the presence of membranes. Consistent with these findings, hydrodynamic analysis of endogenous ESCRT-0 indicates that it exists largely as a heterotetrameric complex of its two subunits. Based on these data, we present a revised model for ESCRT-0 function in cargo recruitment and concentration at the endosome.

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Section: Discussionmentioning

confidence: 99%

ESCRT-0 Assembles as a Heterotetrameric Complex on Membranes and Binds Multiple Ubiquitinylated Cargoes Simultaneously

Mayers¹,

Fyfe

Schuh³

et al. 2011

Journal of Biological Chemistry

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“…Inactivation of individual UBDs causes specific defects in the sorting of Ub-cargo into MVBs without compromising other ESCRT functions such as maintenance of endosomal morphology and recycling of proteins between endosomes and the trans-Golgi network (TGN). However, when multiple UBDs are lost, or UBD loss occurs in the context of mutations that weaken the ability of ESCRTs to bind one another, the ESCRT pathway is more broadly compromised [4][5][6]. This suggests that UBDs have an intimate role in controlling ESCRT activity.…”

Section: Introductionmentioning

confidence: 99%

Cargo ubiquitination is essential for multivesicular body intralumenal vesicle formation

et al. 2012

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The efficient formation of a variety of transport vesicles is influenced by the presence of cargo, suggesting that cargo itself might have a defining role in vesicle biogenesis. However, definitive in vivo experiments supporting this concept are lacking, as it is difficult to eliminate endogenous cargo. The Endosomal Sorting Complexes Required for Transport (ESCRT) apparatus sorts ubiquitinated membrane proteins into endosomal intralumenal vesicles (ILVs) that accumulate within multivesicular bodies. Here we show that cargo ubiquitination is required for effective recruitment of the ESCRT machinery onto endosomal membranes and for the subsequent formation of ILVs.

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“…This sequential sorting process is orchestrated by the ability of ESCRTs to directly interact with ESCRT-0 binding, activating, and transferring cargo to ESCRT-I, and similarly ESCRT-I to ESCRT-II. The ability of ESCRT-I and -II to form a supercomplex and the lack of a clear necessary role for Ub binding by ESCRT-II suggests ESCRT-I and -II function together rather than individually in sequence, as a secondary Ub-sorting receptor that receives cargo from ESCRT-0 (Amerik et al 2006;Shields et al 2009;Boura et al 2012). The idea that these work in sequence is supported by immunolocalization experiments showing a differential distribution of ESCRT-0 and ESCRT-I, live cell imaging that shows sequential assembly of ESCRTs at the sites of viral budding and cytokinesis, and biochemical studies in yeast mutants where assembly intermediates can be accumulated (Babst et al 2002;Bache et al 2003;Elia et al 2011;Guizetti et al 2011;Jouvenet et al 2011).…”

Section: Sequential Orchestration Of Escrt Functionmentioning

confidence: 99%

“…UBAP1 is one of three alternative Mvb12 orthologs that can be incorporated into the mammalian ESCRT-I heterotetramer, but is the one with the highest Ub-binding activity and appears to be the one most suited for sorting Ub cargo into MVBs (Stefani et al 2011;Agromayor et al 2012). Mutations that inactivate Ub binding by yeast Vps23 and Mvb12 or by mammalian UBAP1 block sorting of Ub cargo suggesting that ESCRT-I also functions as a Ub-sorting receptor (Shields et al 2009;Stefani et al 2011). ESCRT-II binds Ub via its Vps36/Eap45 subunit, which also binds endosomal PtdIns-3P.…”

Section: Ubiquitin-dependent Sorting In Endocytosismentioning

confidence: 99%

Ubiquitin-Dependent Sorting in Endocytosis

Piper

Đikić

Lukacs

2014

Cold Spring Harbor Perspectives in Biology

193

176

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When ubiquitin (Ub) is attached to membrane proteins on the plasma membrane, it directs them through a series of sorting steps that culminate in their delivery to the lumen of the lysosome where they undergo complete proteolysis. Ubiquitin is recognized by a series of complexes that operate at a number of vesicle transport steps. Ubiquitin serves as a sorting signal for internalization at the plasma membrane and is the major signal for incorporation into intraluminal vesicles of multivesicular late endosomes. The sorting machineries that catalyze these steps can bind Ub via a variety of Ub-binding domains. At the same time, many of these complexes are themselves ubiquitinated, thus providing a plethora of potential mechanisms to regulate their activity. Here we provide an overview of how membrane proteins are selected for ubiquitination and deubiquitination within the endocytic pathway and how that ubiquitin signal is interpreted by endocytic sorting machineries. HOW PROTEINS ARE SELECTED FOR UBIQUITINATIONU biquitin (Ub) is covalently attached to substrate proteins by the concerted action of E2-conjugating enzymes and E3 ligases (Varshavsky 2012). Most of the specificity and regulation of ubiquitination is at the level of the E3 ligases, which vastly outnumber the E2-conjugating enzymes. Ubiquitination results from the transfer of Ub, held either by the E2 or in some cases by the E3 as a high-energy thioester bond, onto a substrate protein, typically on the terminal amide of a substrate acceptor lysine side chain. Owing to the intense interest in the ubiquitination system, many of the simple rules and distinctions concerning different types of ligases, their specificity for forming particular polyUb chains, and even the kinds of residues in substrate proteins that can be ubiquitin modified, have been blurred with the discovery of mixed poly-Ub chains, new enzymatic mechanisms for Ub transfer, and ubiquitination of nonlysine residues (Kulathu and Komander 2012;Wenzel and Klevit 2012;McDowell and Philpott 2013). Nonetheless, in basic terms, Ub ligases function as platforms that coordinate substrate recognition with Ub transfer as well as configuring poly-Ub chain topology by the E2-conjugating enzyme. Substrates can be bound directly by the E3 ligase or by adaptor proteins that in turn bind to ligases, and selecEditors:

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ESCRT ubiquitin-binding domains function cooperatively during MVB cargo sorting

Cited by 100 publications

References 38 publications

ESCRT-0 Assembles as a Heterotetrameric Complex on Membranes and Binds Multiple Ubiquitinylated Cargoes Simultaneously

ESCRT-0 Assembles as a Heterotetrameric Complex on Membranes and Binds Multiple Ubiquitinylated Cargoes Simultaneously

Cargo ubiquitination is essential for multivesicular body intralumenal vesicle formation

Ubiquitin-Dependent Sorting in Endocytosis

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