scholarly journals K63-linked ubiquitin chains as a specific signal for protein sorting into the multivesicular body pathway

2009 ◽  
Vol 185 (3) ◽  
pp. 493-502 ◽  
Author(s):  
Elsa Lauwers ◽  
Christophe Jacob ◽  
Bruno André
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Protein Trafficking ◽  
Multivesicular Body ◽  
Adapter Proteins ◽  
Relative Importance ◽  
Specific Signal ◽  
Single Target ◽  
Membrane Protein Trafficking ◽  
Insight Into

A growing number of yeast and mammalian plasma membrane proteins are reported to be modified with K63-linked ubiquitin (Ub) chains. However, the relative importance of this modification versus monoubiquitylation in endocytosis, Golgi to endosome traffic, and sorting into the multivesicular body (MVB) pathway remains unclear. In this study, we show that K63-linked ubiquitylation of the Gap1 permease is essential for its entry into the MVB pathway. Carboxypeptidase S also requires modification with a K63-Ub chain for correct MVB sorting. In contrast, monoubiquitylation of a single target lysine of Gap1 is a sufficient signal for its internalization from the cell surface, and Golgi to endosome transport of the permease requires neither its ubiquitylation nor the Ub-binding GAT (Gga and Tom1) domain of Gga (Golgi localizing, gamma-ear containing, ARF binding) adapter proteins, the latter being crucial for subsequent MVB sorting of the permease. Our data reveal that K63-linked Ub chains act as a specific signal for MVB sorting, providing further insight into the Ub code of membrane protein trafficking.

scholarly journals Kv2.1 cell surface clusters are insertion platforms for ion channel delivery to the plasma membrane

2012 ◽  
Vol 23 (15) ◽  
pp. 2917-2929 ◽  
Author(s):  
Emily Deutsch ◽  
Aubrey V. Weigel ◽  
Elizabeth J. Akin ◽  
Phil Fox ◽  
Gentry Hansen ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Ion Channel ◽  
Protein Trafficking ◽  
Hippocampal Neurons ◽  
Surface Membrane ◽  
Cell Types ◽  
Homogeneous Surface ◽  
Hek Cells ◽  
Membrane Protein Trafficking

Voltage-gated K+ (Kv) channels regulate membrane potential in many cell types. Although the channel surface density and location must be well controlled, little is known about Kv channel delivery and retrieval on the cell surface. The Kv2.1 channel localizes to micron-sized clusters in neurons and transfected human embryonic kidney (HEK) cells, where it is nonconducting. Because Kv2.1 is postulated to be involved in soluble N-ethylmaleimide–sensitive factor attachment protein receptor–mediated membrane fusion, we examined the hypothesis that these surface clusters are specialized platforms involved in membrane protein trafficking. Total internal reflection–based fluorescence recovery after photobleaching studies and quantum dot imaging of single Kv2.1 channels revealed that Kv2.1-containing vesicles deliver cargo at the Kv2.1 surface clusters in both transfected HEK cells and hippocampal neurons. More than 85% of cytoplasmic and recycling Kv2.1 channels was delivered to the cell surface at the cluster perimeter in both cell types. At least 85% of recycling Kv1.4, which, unlike Kv2.1, has a homogeneous surface distribution, is also delivered here. Actin depolymerization resulted in Kv2.1 exocytosis at cluster-free surface membrane. These results indicate that one nonconducting function of Kv2.1 is to form microdomains involved in membrane protein trafficking. This study is the first to identify stable cell surface platforms involved in ion channel trafficking.

scholarly journals Thermoplasmonic nano-rupture of cells reveals Annexin V function in plasma membrane repair

2021 ◽  
Author(s):  
Guillermo S Moreno-Pescador ◽  
Dunya S Aswad ◽  
Christoffer D Florentsen ◽  
Azra Bahadori ◽  
Mohammad R Arastoo ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Photothermal Therapy ◽  
Annexin V ◽  
General Strategy ◽  
Membrane Repair ◽  
Cell Plasma ◽  
Plasma Membrane Repair ◽  
Molecular Components ◽  
Insight Into

Maintaining the integrity of the cell plasma membrane (PM) is critical for the survival of cells. While an efficient PM repair machinery can aid survival of healthy cells by preventing influx of extracellular calcium, it can also constitute an obstacle in drug delivery and photothermal therapy. We show how nanoscopic holes can be applied to the cell surface thus allowing identification of molecular components with a pivotal role in PM repair. Cells are punctured by locally heating gold nanostructures at the cell surface which causes nano-ruptures in cellular PMs. Recruitment of annexin V near the hole is found to locally reshape the ruptured plasma membrane. Experiments using model membranes, containing recombinant annexin V, provide further biophysical insight into the ability of annexin V to reshape edges surrounding a membrane hole. The thermoplasmonic method provides a general strategy to monitor the response to nanoscopic injuries to the cell surface.

scholarly journals A combined EM and proteomic analysis places HIV-1 Vpu at the crossroads of retromer and ESCRT complexes: PTPN23 is a Vpu-cofactor

2021 ◽  
Vol 17 (11) ◽  
pp. e1009409
Author(s):  
Charlotte A. Stoneham ◽  
Simon Langer ◽  
Paul D. De Jesus ◽  
Jacob M. Wozniak ◽  
John Lapek ◽  
...  
Keyword(s):  
Cell Surface ◽  
Protein Trafficking ◽  
Fusion Proteins ◽  
Immune Surveillance ◽  
Accessory Protein ◽  
Wild Type ◽  
Membrane Systems ◽  
Membrane Protein Trafficking ◽  
Cellular Pathways ◽  
Hiv 1

The HIV-1 accessory protein Vpu modulates membrane protein trafficking and degradation to provide evasion of immune surveillance. Targets of Vpu include CD4, HLAs, and BST-2. Several cellular pathways co-opted by Vpu have been identified, but the picture of Vpu’s itinerary and activities within membrane systems remains incomplete. Here, we used fusion proteins of Vpu and the enzyme ascorbate peroxidase (APEX2) to compare the ultrastructural locations and the proximal proteomes of wild type Vpu and Vpu-mutants. The proximity-omes of the proteins correlated with their ultrastructural locations and placed wild type Vpu near both retromer and ESCRT-0 complexes. Hierarchical clustering of protein abundances across the mutants was essential to interpreting the data and identified Vpu degradation-targets including CD4, HLA-C, and SEC12 as well as Vpu-cofactors including HGS, STAM, clathrin, and PTPN23, an ALIX-like protein. The Vpu-directed degradation of BST-2 was supported by STAM and PTPN23 and to a much lesser extent by the retromer subunits Vps35 and SNX3. PTPN23 also supported the Vpu-directed decrease in CD4 at the cell surface. These data suggest that Vpu directs targets from sorting endosomes to degradation at multi-vesicular bodies via ESCRT-0 and PTPN23.

scholarly journals Gibberellin DELLA signaling targets the retromer complex to redirect protein trafficking to the plasma membrane

2018 ◽  
Vol 115 (14) ◽  
pp. 3716-3721 ◽  
Author(s):  
Yuliya Salanenka ◽  
Inge Verstraeten ◽  
Christian Löfke ◽  
Kaori Tabata ◽  
Satoshi Naramoto ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Protein Trafficking ◽  
Molecular Mechanisms ◽  
Sorting Nexin ◽  
Protein Traffic ◽  
Della Proteins ◽  
Retromer Complex ◽  
Ga Signaling ◽  
Multiple Processes

The plant hormone gibberellic acid (GA) is a crucial regulator of growth and development. The main paradigm of GA signaling puts forward transcriptional regulation via the degradation of DELLA transcriptional repressors. GA has also been shown to regulate tropic responses by modulation of the plasma membrane incidence of PIN auxin transporters by an unclear mechanism. Here we uncovered the cellular and molecular mechanisms by which GA redirects protein trafficking and thus regulates cell surface functionality. Photoconvertible reporters revealed that GA balances the protein traffic between the vacuole degradation route and recycling back to the cell surface. Low GA levels promote vacuolar delivery and degradation of multiple cargos, including PIN proteins, whereas high GA levels promote their recycling to the plasma membrane. This GA effect requires components of the retromer complex, such as Sorting Nexin 1 (SNX1) and its interacting, microtubule (MT)-associated protein, the Cytoplasmic Linker-Associated Protein (CLASP1). Accordingly, GA regulates the subcellular distribution of SNX1 and CLASP1, and the intact MT cytoskeleton is essential for the GA effect on trafficking. This GA cellular action occurs through DELLA proteins that regulate the MT and retromer presumably via their interaction partners Prefoldins (PFDs). Our study identified a branching of the GA signaling pathway at the level of DELLA proteins, which, in parallel to regulating transcription, also target by a nontranscriptional mechanism the retromer complex acting at the intersection of the degradation and recycling trafficking routes. By this mechanism, GA can redirect receptors and transporters to the cell surface, thus coregulating multiple processes, including PIN-dependent auxin fluxes during tropic responses.

scholarly journals Normal prion protein trafficking in cultured human erythroblasts

Blood ◽  
2007 ◽  
Vol 110 (13) ◽  
pp. 4518-4525 ◽  
Author(s):  
Rebecca E. Griffiths ◽  
Kate J. Heesom ◽  
David J. Anstee
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Prion Protein ◽  
Protein Trafficking ◽  
Prion Diseases ◽  
Cross Linking ◽  
Red Cell ◽  
Jakob Disease ◽  
Endosomal Pathway ◽  
Tetraspanin Cd63

Normal prion protein (PrPc), an essential substrate for development of prion disease, is widely distributed in hematopoietic cells. Recent evidence that variant Creutzfeldt-Jakob disease can be transmitted by transfusion of red cell preparations has highlighted the need for a greater understanding of the biology of PrPc in blood and blood-forming tissues. Here, we show that in contrast to another glycosylphosphoinositol-anchored protein CD59, PrPc at the cell surface of cultured human erythroblasts is rapidly internalized through the endosomal pathway, where it colocalizes with the tetraspanin CD63. In the plasma membrane, PrPc colocalizes with the tetraspanin CD81. Cross-linking with anti-PrPc or anti-CD81 causes clustering of PrPc and CD81, suggesting they can share the same microdomain. These data are consistent with a role for tetraspanin-enriched microdomains in trafficking of PrPc. These results, when taken together with recent evidence that exosomes released from cells as a result of endosomal-mediated recycling to the plasma membrane contain prion infectivity, provide a pathway for the propagation of prion diseases.

scholarly journals Knockout of Syntaxin-4 in 3T3-L1 adipocytes reveals new insight into GLUT4 trafficking and adiponectin secretion

2021 ◽  
Author(s):  
Hannah L. Black ◽  
Rachel Livingstone ◽  
Cynthia C. Mastick ◽  
Mohammed Al Tobi ◽  
Holly Taylor ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Glucose Transport ◽  
Metabolic Regulation ◽  
Glucose Transporter ◽  
Ectopic Expression ◽  
Glut4 Translocation ◽  
Syntaxin 4 ◽  
Rate Limiting ◽  
Insight Into

Adipocytes are key to metabolic regulation, exhibiting insulin-stimulated glucose transport which is underpinned by the insulin-stimulated delivery of glucose transporter-4 (GLUT4)- containing vesicles to the plasma membrane where they dock and fuse increasing cell surface GLUT4 levels. Adipocytokines such as adiponectin are secreted via a similar mechanism. We used genome editing to knockout Syntaxin-4 a protein reported to mediate GLUT4-vesicle fusion with the plasma membrane in 3T3-L1 adipocytes. Syntaxin-4 knockout reduced insulin-stimulated glucose transport and adiponectin secretion by ∼50% and reduced GLUT4 levels. Ectopic expression of HA-GLUT4-GFP showed that Syntaxin-4 knockout cells retain significant GLUT4 translocation capacity demonstrating that Syntaxin-4 is dispensable for insulin-stimulated GLUT4 translocation. Analysis of recycling kinetics revealed only a modest reduction in the exocytic rate of GLUT4 in knockout cells, and little effect on endocytosis. These analyses demonstrate that Syntaxin-4 is not always rate limiting for GLUT4 delivery to the cell surface. In sum, we show that Syntaxin-4 knockout results in reduced insulin-stimulated glucose transport, depletion of cellular GLUT4 levels and inhibition of adiponectin secretion but has only modest effects on the translocation capacity of the cells.

scholarly journals Uni-directional ciliary membrane protein trafficking by a cytoplasmic retrograde IFT motor and ciliary ectosome shedding

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Muqing Cao ◽  
Jue Ning ◽  
Carmen I Hernandez-Lara ◽  
Olivier Belzile ◽  
Qian Wang ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Protein Trafficking ◽  
Protein Composition ◽  
Cytoplasmic Dynein ◽  
Cytoplasmic Microtubule ◽  
Dynamic Regulation ◽  
Ciliary Membrane ◽  
Membrane Protein Trafficking

The role of the primary cilium in key signaling pathways depends on dynamic regulation of ciliary membrane protein composition, yet we know little about the motors or membrane events that regulate ciliary membrane protein trafficking in existing organelles. Recently, we showed that cilium-generated signaling in Chlamydomonas induced rapid, anterograde IFT-independent, cytoplasmic microtubule-dependent redistribution of the membrane polypeptide, SAG1-C65, from the plasma membrane to the periciliary region and the ciliary membrane. Here, we report that the retrograde IFT motor, cytoplasmic dynein 1b, is required in the cytoplasm for this rapid redistribution. Furthermore, signaling-induced trafficking of SAG1-C65 into cilia is unidirectional and the entire complement of cellular SAG1-C65 is shed during signaling and can be recovered in the form of ciliary ectosomes that retain signal-inducing activity. Thus, during signaling, cells regulate ciliary membrane protein composition through cytoplasmic action of the retrograde IFT motor and shedding of ciliary ectosomes.

Versatile role of the yeast ubiquitin ligase Rsp5p in intracellular trafficking

2008 ◽  
Vol 36 (5) ◽  
pp. 791-796 ◽  
Author(s):  
Naima Belgareh-Touzé ◽  
Sébastien Léon ◽  
Zoi Erpapazoglou ◽  
Marta Stawiecka-Mirota ◽  
Danièle Urban-Grimal ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Ubiquitin Ligase ◽  
Protein Trafficking ◽  
Adaptor Proteins ◽  
Binding Domains ◽  
Independent Functions ◽  
Membrane Protein Trafficking ◽  
Ubiquitin Ligase E3

The ubiquitin ligase (E3) Rsp5p is the only member of the Nedd (neural-precursor-cell-expressed, developmentally down-regulated) 4 family of E3s present in yeast. Rsp5p has several proteasome-independent functions in membrane protein trafficking, including a role in the ubiquitination of most plasma membrane proteins, leading to their endocytosis. Rsp5p is also required for the ubiquitination of endosomal proteins, leading to their sorting to the internal vesicles of MVBs (multivesicular bodies). Rsp5p catalyses the attachment of non-conventional ubiquitin chains, linked through ubiquitin Lys-63, to some endocytic and MVB cargoes. This modification appears to be required for efficient sorting, possibly because these chains have a greater affinity for the ubiquitin-binding domains present within endocytic or MVB sorting complexes. The mechanisms involved in the recognition of plasma membrane and MVB substrates by Rsp5p remain unclear. A subset of Rsp5/Nedd4 substrates have a ‘PY motif’ and are recognized directly by the WW (Trp-Trp) domains of Rsp5p. Most Rsp5p substrates do not carry PY motifs, but some may depend on PY-containing proteins for their ubiquitination by Rsp5p, consistent with the latter's acting as specificity factors or adaptors. As in other ubiquitin-conjugating systems, these adaptors are also Rsp5p substrates and undergo ubiquitin-dependent trafficking. In the present review, we discuss recent examples illustrating the role of Rsp5p in membrane protein trafficking and providing new insights into the regulation of this E3 by adaptor proteins.

Sign in / Sign up

Export Citation Format

Share Document