The Experts below are selected from a list of 11793 Experts worldwide ranked by ideXlab platform
Beverly Wendland - One of the best experts on this subject based on the ideXlab platform.
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ptdins 3 5 p2 is required for delivery of endocytic cargo into the Multivesicular Body
Traffic, 2003Co-Authors: Jonathan D Shaw, Hiroko Hama, Farrokh Sohrabi, Daryll B Dewald, Beverly WendlandAbstract:The endocytic pathway transports cargo from the plasma membrane to early endosomes, where certain cargoes are sorted to the late endosome/Multivesicular Body. Biosynthetic cargo destined for the lysosome is also trafficked through the Multivesicular Body. Once delivered to the Multivesicular Body, cargo destined for the interior of the lysosome is selectively sorted into vesicles that bud into the lumen of the Multivesicular Body. These vesicles are released into the lumen of the lysosome upon the fusion of the Multivesicular Body and lysosomal limiting membranes. The yeast protein Fab1, which catalyzes the production of phosphatidylinositol (3,5) bisphosphate [PtdIns(3,5)P2], is necessary for proper sorting of biosynthetic cargo in the Multivesicular Body. Utilizing an endocytosis screen, we isolated a novel allele of FAB1 that contains a point mutation in the lipid kinase domain. Characterization of this allele revealed reduced PtdIns(3,5)P2 production, altered vacuole morphology, and biosynthetic protein sorting defects. We also found that endocytosis of the plasma membrane protein Ste3 is partially blocked downstream of the internalization step, and that delivery of the dye FM4-64 to the vacuole is delayed in fab1 mutants. Additionally, Ste3 is not efficiently sorted into Multivesicular Body vesicles in fab1 mutants and instead localizes to the vacuolar limiting membrane. These data show that PtdIns(3,5)P2 is necessary for proper trafficking and sorting of endocytic cargo through the late endosome/Multivesicular Body.
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endosome associated complex escrt ii recruits transport machinery for protein sorting at the Multivesicular Body
Developmental Cell, 2002Co-Authors: Markus Babst, David J Katzmann, William B Snyder, Beverly WendlandAbstract:Abstract Sorting of ubiquitinated endosomal membrane proteins into the MVB pathway is executed by the class E Vps protein complexes ESCRT-I, -II, and -III, and the AAA-type ATPase Vps4. This study characterizes ESCRT-II, a soluble ∼155 kDa protein complex formed by the class E Vps proteins Vps22, Vps25, and Vps36. This protein complex transiently associates with the endosomal membrane and thereby initiates the formation of ESCRT-III, a membrane-associated protein complex that functions immediately downstream of ESCRT-II during sorting of MVB cargo. ESCRT-II in turn functions downstream of ESCRT-I, a protein complex that binds to ubiquitinated endosomal cargo. We propose that the ESCRT complexes perform a coordinated cascade of events to select and sort MVB cargoes for delivery to the lumen of the vacuole/lysosome.
Greg Odorizzi - One of the best experts on this subject based on the ideXlab platform.
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bro1 directly stimulates vps4 activity to promote intralumenal vesicle formation during Multivesicular Body biogenesis
bioRxiv, 2020Co-Authors: Chunche Tseng, Shirley Dean, Brian A Davies, Ishara F Azmi, Johanna A Payne, Robert C Piper, Natalya Pashkova, Jennifer Staffenhagen, Matthew West, Greg OdorizziAbstract:Abstract Endosomal sorting complexes required for transport (ESCRT-0, -I, -II, -III) form intralumenal vesicles (ILVs) during the conversion of endosomes to Multivesicular bodies (MVBs). The assembly and disassembly of an ESCRT-III polymer facilitates membrane remodeling and scission during this process. The ESCRT-III-associated protein Bro1 (the yeast homolog of mammalian proteins ALIX and HD-PTP) promotes ESCRT-III assembly and inhibits disassembly of ESCRT-III filaments by impeding Vps4, a AAA-ATPase that dismantles ESCRT-III polymers. Here we show that the evolutionarily conserved “V domain” of Bro1-family proteins directly stimulate Vps4 ATPase activity and this activity is enhanced by interaction with ubiquitin. Surprisingly, a carboxyl-terminal fragment of Bro1 containing the V domain supports ILV formation but not sorting of cargo into ILVs, revealing that these two processes can be uncoupled. These studies implicate Bro1 as a factor coordinating cargo sorting with direct regulation of Vps4 to modulate ESCRT-III driven ILV formation during MVB biogenesis.
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endosomal na k h exchanger nhx1 vps44 functions independently and downstream of Multivesicular Body formation
Journal of Biological Chemistry, 2011Co-Authors: Laura Kallay, Christopher L Brett, Deepali N Tukaye, Megan Wemmer, Anthony Chyou, Greg OdorizziAbstract:The Multivesicular Body (MVB) is an endosomal intermediate containing intralumenal vesicles destined for membrane protein degradation in the lysosome. In Saccharomyces cerevisiae, the MVB pathway is composed of 17 evolutionarily conserved ESCRT (endosomal sorting complex required for transport) genes grouped by their vacuole protein sorting Class E mutant phenotypes. Only one integral membrane protein, the endosomal Na+ (K+)/H+ exchanger Nhx1/Vps44, has been assigned to this class, but its role in the MVB pathway has not been directly tested. Herein, we first evaluated the link between Nhx1 and the ESCRT proteins and then used an unbiased phenomics approach to probe the cellular role of Nhx1. Select ESCRT mutants (vps36Δ, vps20Δ, snf7Δ, and bro1Δ) with defects in cargo packaging and intralumenal vesicle formation shared multiple growth phenotypes with nhx1Δ. However, analysis of cellular trafficking and ultrastructural examination by electron microscopy revealed that nhx1Δ cells retain the ability to sort cargo into intralumenal vesicles. In addition, we excluded a role for Nhx1 in Snf7/Bro1-mediated cargo deubiquitylation and Rim101 response to pH stress. Genetic epistasis experiments provided evidence that NHX1 and ESCRT genes function in parallel. A genome-wide screen for single gene deletion mutants that phenocopy nhx1Δ yielded a limited gene set enriched for endosome fusion function, including Rab signaling and actin cytoskeleton reorganization. In light of these findings and the absence of the so-called Class E compartment in nhx1Δ, we eliminated a requirement for Nhx1 in MVB formation and suggest an alternative post-ESCRT role in endosomal membrane fusion.
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A concentric circle model of Multivesicular Body cargo sorting
EMBO Reports, 2007Co-Authors: David P. Nickerson, M. R G Russell, Greg OdorizziAbstract:Targeting of ubiquitylated transmembrane proteins into luminal vesicles of endosomal Multivesicular bodies (MVBs) depends on their recognition by endosomal sorting complexes required for transport (ESCRTs), which are also required for MVB vesicle formation. The model originally proposed for how ESCRTs function succinctly summarizes much of the protein-protein interaction and genetic data but oversimplifies the coordination of cargo recognition and cannot explain why ESCRTs are required for the budding of MVB vesicles. Recent structural and functional studies of ESCRT complexes suggest an alternative model that might direct the next series of breakthroughs in understanding protein sorting through the MVB pathway.
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concept a concentric circle model of Multivesicular Body cargo sorting
2007Co-Authors: David P. Nickerson, Greg OdorizziAbstract:Targeting of ubiquitylated transmembrane proteins into luminal vesicles of endosomal Multivesicular bodies (MVBs) depends on their recognition by endosomal sorting complexes required for transport (ESCRTs), which are also required for MVB vesicle formation. The model originally proposed for how ESCRTs function succinctly summarizes much of the protein‐protein interaction and genetic data but oversimplifies the coordination of cargo recognition and cannot explain why ESCRTs are required for the budding of MVB vesicles. Recent structural and functional studies of ESCRT complexes suggest an alternative model that might direct the next series of breakthroughs in understanding protein sorting through the MVB pathway.
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bro1 coordinates deubiquitination in the Multivesicular Body pathway by recruiting doa4 to endosomes
Journal of Cell Biology, 2004Co-Authors: Natalie Luhtala, Greg OdorizziAbstract:Ubiquitination directs the sorting of cell surface receptors and other integral membrane proteins into the Multivesicular Body (MVB) pathway. Cargo proteins are subsequently deubiquitinated before their enclosure within MVB vesicles. In Saccharomyces cerevisiae, Bro1 functions at a late step of MVB sorting and is required for cargo protein deubiquitination. We show that the loss of Bro1 function is suppressed by the overexpression of DOA4, which encodes the ubiquitin thiolesterase required for the removal of ubiquitin from MVB cargoes. Overexpression of DOA4 restores cargo protein deubiquitination and sorting via the MVB pathway and reverses the abnormal endosomal morphology typical of bro1 mutant cells, resulting in the restoration of Multivesicular endosomes. We further demonstrate that Doa4 interacts with Bro1 on endosomal membranes and that the recruitment of Doa4 to endosomes requires Bro1. Thus, our results point to a key role for Bro1 in coordinating the timing and location of deubiquitination by Doa4 in the MVB pathway.
Francois Letourneur - One of the best experts on this subject based on the ideXlab platform.
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ent5p is required with ent3p and vps27p for ubiquitin dependent protein sorting into the Multivesicular Body
Molecular Biology of the Cell, 2004Co-Authors: Anne Eugster, Eveisabelle Pecheur, Fabrice Michel, Barbara Winsor, Francois Letourneur, Sylvie FriantAbstract:At the late endosomes, cargoes destined for the interior of the vacuole are sorted into invaginating vesicles of the Multivesicular Body. Both PtdIns(3,5)P(2) and ubiquitin are necessary for proper sorting of some of these cargoes. We show that Ent5p, a yeast protein of the epsin family homologous to Ent3p, localizes to endosomes and specifically binds to PtdIns(3,5)P(2) via its ENTH domain. In cells lacking Ent3p and Ent5p, ubiquitin-dependent sorting of biosynthetic and endocytic cargo into the Multivesicular Body is disrupted, whereas other trafficking routes to the vacuole are not affected. Ent3p and Ent5p are associated with Vps27p, a FYVE domain containing protein that interacts with ubiquitinated cargoes and is required for protein sorting into the Multivesicular Body. Therefore, Ent3p and Ent5p are the first proteins shown to be connectors between PtdIns(3,5)P(2)- and the Vps27p-ubiquitin-driven sorting machinery at the Multivesicular Body.
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ent3p is a ptdins 3 5 p2 effector required for protein sorting to the Multivesicular Body
Developmental Cell, 2003Co-Authors: Sylvie Friant, Anne Eugster, Eveisabelle Pecheur, Fabrice Michel, Yaya Lefkir, Delphine Nourrisson, Francois LetourneurAbstract:Abstract PtdIns(3,5)P 2 is required for cargo-selective sorting to the vacuolar lumen via the Multivesicular Body (MVB). Here we show that Ent3p, a yeast epsin N-terminal homology (ENTH) domain-containing protein, is a specific PtdIns(3,5)P 2 effector localized to endosomes. The ENTH domain of Ent3p is essential for its PtdIns(3,5)P 2 binding activity and for its membrane interaction in vitro and in vivo. Ent3p is required for protein sorting into the MVB but not for the internalization step of endocytosis. Ent3p is associated with clathrin and is necessary for normal actin cytoskeleton organization. Our results show that Ent3p is required for protein sorting into intralumenal vesicles of the MVB through PtdIns(3,5)P 2 binding via its ENTH domain.
Sylvie Friant - One of the best experts on this subject based on the ideXlab platform.
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ent5p is required with ent3p and vps27p for ubiquitin dependent protein sorting into the Multivesicular Body
Molecular Biology of the Cell, 2004Co-Authors: Anne Eugster, Eveisabelle Pecheur, Fabrice Michel, Barbara Winsor, Francois Letourneur, Sylvie FriantAbstract:At the late endosomes, cargoes destined for the interior of the vacuole are sorted into invaginating vesicles of the Multivesicular Body. Both PtdIns(3,5)P(2) and ubiquitin are necessary for proper sorting of some of these cargoes. We show that Ent5p, a yeast protein of the epsin family homologous to Ent3p, localizes to endosomes and specifically binds to PtdIns(3,5)P(2) via its ENTH domain. In cells lacking Ent3p and Ent5p, ubiquitin-dependent sorting of biosynthetic and endocytic cargo into the Multivesicular Body is disrupted, whereas other trafficking routes to the vacuole are not affected. Ent3p and Ent5p are associated with Vps27p, a FYVE domain containing protein that interacts with ubiquitinated cargoes and is required for protein sorting into the Multivesicular Body. Therefore, Ent3p and Ent5p are the first proteins shown to be connectors between PtdIns(3,5)P(2)- and the Vps27p-ubiquitin-driven sorting machinery at the Multivesicular Body.
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ent3p is a ptdins 3 5 p2 effector required for protein sorting to the Multivesicular Body
Developmental Cell, 2003Co-Authors: Sylvie Friant, Anne Eugster, Eveisabelle Pecheur, Fabrice Michel, Yaya Lefkir, Delphine Nourrisson, Francois LetourneurAbstract:Abstract PtdIns(3,5)P 2 is required for cargo-selective sorting to the vacuolar lumen via the Multivesicular Body (MVB). Here we show that Ent3p, a yeast epsin N-terminal homology (ENTH) domain-containing protein, is a specific PtdIns(3,5)P 2 effector localized to endosomes. The ENTH domain of Ent3p is essential for its PtdIns(3,5)P 2 binding activity and for its membrane interaction in vitro and in vivo. Ent3p is required for protein sorting into the MVB but not for the internalization step of endocytosis. Ent3p is associated with clathrin and is necessary for normal actin cytoskeleton organization. Our results show that Ent3p is required for protein sorting into intralumenal vesicles of the MVB through PtdIns(3,5)P 2 binding via its ENTH domain.
Paul Ahlquist - One of the best experts on this subject based on the ideXlab platform.
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intersection of the Multivesicular Body pathway and lipid homeostasis in rna replication by a positive strand rna virus
Journal of Virology, 2011Co-Authors: Xiaofeng Wang, Arturo Diaz, Brandi Gancarz, Johan Den A Boon, Paul AhlquistAbstract:Like many positive-strand RNA viruses, brome mosaic virus (BMV) RNA replication occurs in membrane-invaginated vesicular compartments. BMV RNA replication compartments show parallels with membrane-enveloped, budding retrovirus virions, whose release depends on the cellular Multivesicular Body (MVB) sorting pathway. BMV RNA replication compartments are not released from their parent membranes, but might depend on MVB functions for membrane invagination. Prior results show that BMV RNA replication is severely inhibited by deletion of the crucial MVB gene DOA4 or BRO1. We report here that involvement of DOA4 and BRO1 in BMV RNA replication is not dependent on the MVB pathway's membrane-shaping functions but rather is due to their roles in recycling ubiquitin from MVB cargos. We show that deleting DOA4 or BRO1 inhibits the ubiquitination- and proteasome-dependent activation of homologous transcription factors Mga2p and Spt23p, which regulate many lipid metabolism genes, including the fatty acid desaturase gene OLE1, which is essential for BMV RNA replication. However, Mga2p processing and BMV RNA replication are restored by supplementing free ubiquitin, which is depleted in doa4Δ and bro1Δ cells. The results identify Mga2p and Spt23p processing and lipid regulation as sensitive targets of ubiquitin depletion and correctly predict multiple effects of modulating additional host genes RFU1, UBP6, and UFD3. Our results also show that BMV RNA replication depends on additional Mga2p-regulated genes likely involved in lipid metabolism beyond OLE1. Among other points, these findings show the potential for blocking viral RNA replication by modulating lipid synthesis at multiple levels.
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involvement of host cellular Multivesicular Body functions in hepatitis b virus budding
Proceedings of the National Academy of Sciences of the United States of America, 2007Co-Authors: Tokiko Watanabe, Ericka Sorensen, Akira Naito, Meghan Schott, Paul AhlquistAbstract:Hepatitis B virus (HBV) is a major human pathogen that chronically infects ≈350 million people, causing liver disease and liver cancer. HBV virions bud into an endoplasmic reticulum (ER)-associated intracellular compartment, but the mechanisms of HBV assembly, budding, and release remain poorly understood. Budding of retroviruses and some other enveloped RNA viruses from plasma membranes requires host functions involved in protein sorting into late endosomal Multivesicular bodies (MVBs). To determine whether budding of DNA-containing HBV virions at intracellular membranes also involves MVB functions, we used immunofluorescence to show that, in human hepatoma cells, HBV envelope protein colocalizes with MVB proteins AIP1/ALIX and VPS4B. We also found that a dominant negative (DN) AIP1 mutant inhibited production and/or release of enveloped virions without significant effects on intracellular nucleocapsid formation, whereas DN VPS4B inhibited both nucleocapsid production and budding. By contrast, DN AIP1 and VPS4 had no effect on the efficiency of release of enveloped, nucleocapsid-lacking HBV subviral particles, which are produced in vast excess over virions, and dramatically increased the release of unenveloped, naked nucleocapsids by an apparently nonlytic route. Thus, host MVB functions are required for efficient budding and release of enveloped HBV virions and may be a valuable target for HBV control. Moreover, HBV enveloped virions, enveloped subviral particles, and unenveloped nucleocapsids are all released by distinct pathways with separate host factor requirements.
