Hse1, a Component of the Yeast Hrs-STAM Ubiquitin-sorting Complex, Associates with Ubiquitin Peptidases and a Ligase to Control Sorting Efficiency into Multivesicular Bodies

Ren, Ji‐Chang; Kee, Younghoon; Huibregtse, Jon M.; Piper, Robert C.

doi:10.1091/mbc.e06-06-0557

Cited by 68 publications

(81 citation statements)

References 80 publications

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“…However, previous studies indicate that both Hrs and STAM can associate with several other proteins, potentially accounting for the unexpectedly large mass of the complex. STAM has been shown to interact with multiple deubiquitinating enzymes and a ubiquitin ligase, which may help to regulate the sorting efficiency of cargoes into MVEs (31,32). Hrs has been implicated in the recruitment of clathrin to MVEs, which likely functions to retain cargoes within microdomains of early endosomes (14,33).…”

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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“…Indeed, one member of the Nedd4 family stably associates with Tsg101, a subunit of ESCRT-I (Medina et al, 2005), indicating a physical link exists between the E3 Ub ligase and core MVB sorting components. Recently, Rsp5 was found to bind directly to Hse1, a subunit of the ESCRT-0 complex (Ren et al, 2006), which might explain how Sna3 bound to Rsp5 gains accesses the class E Vps/ESCRT MVB sorting machinery.…”

Section: Discussionmentioning

confidence: 99%

Direct Binding to Rsp5 Mediates Ubiquitin-independent Sorting of Sna3 via the Multivesicular Body Pathway

McNatt

McKittrick

West

et al. 2007

MBoC

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The sorting of most integral membrane proteins into the lumenal vesicles of multivesicular bodies (MVBs) is dependent on the attachment of ubiquitin (Ub) to their cytosolic domains. However, Ub is not required for sorting of Sna3, an MVB vesicle cargo protein in yeast. We show that Sna3 circumvents Ub-mediated recognition by interacting directly with Rsp5, an E3 Ub ligase that catalyzes monoubiquitination of MVB vesicle cargoes. The PPAY motif in the C-terminal cytosolic domain of Sna3 binds the WW domains in Rsp5, and Sna3 is polyubiquitinated as a consequence of this association. However, Ub does not appear to be required for transport of Sna3 via the MVB pathway because its sorting occurs under conditions in which its ubiquitination is impaired. Consistent with Ub-independent function of the MVB pathway, we show by electron microscopy that the formation of MVB vesicles does not require Rsp5 E3 ligase activity. However, cells expressing a catalytically disabled form of Rsp5 have a greater frequency of smaller MVB vesicles compared with the relatively broad distribution of vesicles seen in MVBs of wild-type cells, suggesting that the formation of MVB vesicles is influenced by Rsp5-mediated ubiquitination. INTRODUCTIONMultivesicular bodies (MVBs) are late endosomes containing lumenal vesicles that are formed by invagination of the endosomal membrane. The lumenal MVB vesicles are delivered into the hydrolytic interior of the lysosome upon fusion of the limiting MVB membrane with the lysosomal membrane (van Deurs et al., 1995;Futter et al., 1996;Mullock et al., 1998). Many cell-surface receptors that undergo down-regulation from the plasma membrane are sorted into MVB vesicles en route to being degraded in the lysosome, including members of the receptor tyrosine kinase family in metazoans and G protein-coupled receptors in the budding yeast Saccharomyces cerevisiae. In yeast, many biosynthetic proteins are also sorted into MVB vesicles during their transport from the Golgi to the vacuole, the functional equivalent of the lysosome (reviewed in Hicke and Dunn, 2003).Most integral membrane proteins require the attachment of a single ubiquitin (Ub) to their cytosolic domains in order to be sorted into the MVB pathway (Katzmann et al., 2001;Reggiori and Pelham, 2001;Urbanowski and Piper, 2001). Sorting of monoubiquitinated MVB cargoes is mediated by a highly conserved machinery comprised of class E Vps proteins, many of which assemble into distinct endosomal sorting complexes required for transport (ESCRTs) that contain Ub-binding domains (reviewed in Hurley and Emr, 2006). In addition to Ub-mediated cargo recognition, class E Vps proteins are generally required for MVB vesicle budding because cells in which class E Vps proteins are disrupted contain malformed late endosomes that lack lumenal vesicles (Rieder et al., 1996;Odorizzi et al., 1998;Doyotte et al., 2005).The class E Vps/ESCRT machinery is also required for the budding of many nonlytic enveloped viruses from infected host cells, which is topologically equival...

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“…However, AMSH may also function at an early step in the MVB sorting pathway via additional interactions with STAM and clathrin; AMSH may provide an editing function to precisely regulate the disposition of cargo when AMSH is recognized by the MVB sorting machinery (Bowers et al 2006, McCullough et al 2004. Similarly, in yeast the STAM homolog Hse1 associates with both the HECT Ub ligase Rsp5 and DUbs to regulate the efficiency of cargo sorting (Ren et al 2007). This earlier role for DUb enzymes suggests that ubiquitination is a dynamic process wherein MVB cargoes likely undergo multiple rounds of ubiquitination/ deubiquitination while they travel toward the MVB interior.…”

Section: Lumen Ubiquitin As a Sorting Signalmentioning

confidence: 99%

Biogenesis and Function of Multivesicular Bodies

Piper

Katzmann

2007

Annu. Rev. Cell Dev. Biol.

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648

660

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The two major cellular sites for membrane protein degradation are the proteasome and the lysosome. Ubiquitin attachment is a sorting signal for both degradation routes. For lysosomal degradation, ubiquitination triggers the sorting of cargo proteins into the lumen of late endosomal multivesicular bodies (MVBs)/endosomes. MVB formation occurs when a portion of the limiting membrane of an endosome invaginates and buds into its own lumen. Intralumenal vesicles are degraded when MVBs fuse to lysosomes. The proper delivery of proteins to the MVB interior relies on specific ubiquitination of cargo, recognition and sorting of ubiquitinated cargo to endosomal subdomains, and the formation and scission of cargo-filled intralumenal vesicles. Over the past five years, a number of proteins that may directly participate in these aspects of MVB function and biogenesis have been identified. However, major questions remain as to exactly what these proteins do at the molecular level and how they may accomplish these tasks.

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Hse1, a Component of the Yeast Hrs-STAM Ubiquitin-sorting Complex, Associates with Ubiquitin Peptidases and a Ligase to Control Sorting Efficiency into Multivesicular Bodies

Cited by 68 publications

References 80 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

Direct Binding to Rsp5 Mediates Ubiquitin-independent Sorting of Sna3 via the Multivesicular Body Pathway

Biogenesis and Function of Multivesicular Bodies

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