Golgi complex–plasma membrane trafficking directed by an autonomous, tribasic Golgi export signal

Although numerous linear motifs that direct protein trafficking within cells have been identified, there are few examples of linear sorting signals mediating directed export of membrane proteins from the Golgi complex to the plasma membrane. The reovirus fusion-associated small transmembrane proteins are simple, single-pass transmembrane proteins that traffic through the endoplasmic reticulum–Golgi pathway to the plasma membrane, where they induce cell–cell membrane fusion. Here we show that a membrane-proximal, polybasic motif (PBM) in the cytosolic tail of p14 is essential for efficient export of p14 from the Golgi complex to the plasma membrane. Extensive mutagenic analysis reveals that the number, but not the identity or position, of basic residues present in the PBM dictates p14 export from the Golgi complex, with a minimum of three basic residues required for efficient Golgi export. Results further indicate that the tribasic motif does not affect plasma membrane retention of p14. Furthermore, introduction of the tribasic motif into a Golgi-localized, chimeric ERGIC-53 protein directs export from the Golgi complex to the plasma membrane. The p14 PBM is the first example of an autonomous, tribasic signal required for Golgi export to the plasma membrane.

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Rab proteins of the endoplasmic reticulum: functions and interactors

Biochemical Society Transactions ◽

10.1042/bst20120158 ◽

2012 ◽

Vol 40 (6) ◽

pp. 1426-1432 ◽

Cited By ~ 30

Author(s):

Carolina Ortiz Sandoval ◽

Thomas Simmen

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Golgi Complex ◽

Membrane Trafficking ◽

Small Gtpases ◽

Rab Proteins ◽

Retrograde Trafficking ◽

Rab Family ◽

Intermediate Compartment

Whereas most of what we know today about the Ras-related small GTPases of the Rab family stems from observations made on Golgi complex, endosome and plasma membrane trafficking, a subset of Rabs localizes in part or predominantly to the ER (endoplasmic reticulum). Here, Rabs such as Rab1, Rab2, Rab6 and Rab33 can regulate the anterograde and retrograde trafficking of vesicles between the Golgi complex, the ERGIC (ER–Golgi intermediate compartment) and the ER itself. However, among the ER-associated Rabs, some Rabs appear to perform roles not directly related to trafficking: these Rabs (e.g. Rab32 or Rab24) could aid proteins of the atlastin and reticulon families in determining the extent and direction of ER tubulation. In so doing, these Rabs regulate not only ER contacts with other organelles such as mitochondria, but also the formation of autophagosomes.

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Plasma membrane domains enriched in cortical endoplasmic reticulum function as membrane protein trafficking hubs

Molecular Biology of the Cell ◽

10.1091/mbc.e12-12-0895 ◽

2013 ◽

Vol 24 (17) ◽

pp. 2703-2713 ◽

Cited By ~ 24

Author(s):

Philip D. Fox ◽

Christopher J. Haberkorn ◽

Aubrey V. Weigel ◽

Jenny L. Higgins ◽

Elizabeth J. Akin ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Membrane Protein ◽

Membrane Trafficking ◽

Protein Trafficking ◽

Mammalian Cells ◽

Transmembrane Protein ◽

Surface Proteins ◽

Hek 293 ◽

Cortical Endoplasmic Reticulum

In mammalian cells, the cortical endoplasmic reticulum (cER) is a network of tubules and cisterns that lie in close apposition to the plasma membrane (PM). We provide evidence that PM domains enriched in underlying cER function as trafficking hubs for insertion and removal of PM proteins in HEK 293 cells. By simultaneously visualizing cER and various transmembrane protein cargoes with total internal reflectance fluorescence microscopy, we demonstrate that the majority of exocytotic delivery events for a recycled membrane protein or for a membrane protein being delivered to the PM for the first time occur at regions enriched in cER. Likewise, we observed recurring clathrin clusters and functional endocytosis of PM proteins preferentially at the cER-enriched regions. Thus the cER network serves to organize the molecular machinery for both insertion and removal of cell surface proteins, highlighting a novel role for these unique cellular microdomains in membrane trafficking.

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Stromuling when stressed!

Acta Societatis Botanicorum Poloniae ◽

10.5586/asbp.2014.050 ◽

2014 ◽

Vol 83 (4) ◽

pp. 325-329 ◽

Cited By ~ 4

Author(s):

Björn Krenz ◽

Tai Wei Guo ◽

Tatjana Kleinow

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Golgi Complex ◽

Plant Virus ◽

Complex Plasma ◽

Envelope Membranes

Stromules are stroma-filled tubules, extruding from the plastid and surrounded by both envelope membranes, but so far, stromules remain enigmatic structures and their function unknown. Stromules can interconnect plastids and have been found to associate with the nucleus, endoplasmic reticulum, Golgi complex, plasma membrane, mitochondria and peroxisomes. This minireview briefly summarizes markers to visualize stromules, inducers of stromules and provides new data about plant virus induced stromules.

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Prohibitin: A Novel Molecular Player in KDEL Receptor Signalling

BioMed Research International ◽

10.1155/2015/319454 ◽

2015 ◽

Vol 2015 ◽

pp. 1-13 ◽

Cited By ~ 8

Author(s):

Monica Giannotta ◽

Giorgia Fragassi ◽

Antonio Tamburro ◽

Capone Vanessa ◽

Alberto Luini ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Golgi Complex ◽

Membrane Trafficking ◽

Transmembrane Domain ◽

Cell Cycle Control ◽

Retrograde Transport ◽

Multifunctional Protein ◽

G Protein Coupled ◽

Kdel Receptor ◽

Prohibitin 1

The KDEL receptor (KDELR) is a seven-transmembrane-domain protein involved in retrograde transport of protein chaperones from the Golgi complex to the endoplasmic reticulum. Our recent findings have shown that the Golgi-localised KDELR acts as a functional G-protein-coupled receptor by binding to and activating Gs and Gq. These G proteins induce activation of PKA and Src and regulate retrograde and anterograde Golgi trafficking. Here we used an integrated coimmunoprecipitation and mass spectrometry approach to identify prohibitin-1 (PHB) as a KDELR interactor. PHB is a multifunctional protein that is involved in signal transduction, cell-cycle control, and stabilisation of mitochondrial proteins. We provide evidence that depletion of PHB induces intense membrane-trafficking activity at the ER–Golgi interface, as revealed by formation of GM130-positive Golgi tubules, and recruitment of p115,β-COP, and GBF1 to the Golgi complex. There is also massive recruitment of SEC31 to endoplasmic-reticulum exit sites. Furthermore, absence of PHB decreases the levels of the Golgi-localised KDELR, thus preventing KDELR-dependent activation of Golgi-Src and inhibiting Golgi-to-plasma-membrane transport of VSVG. We propose a model whereby in analogy to previous findings (e.g., the RAS-RAF signalling pathway), PHB can act as a signalling scaffold protein to assist in KDELR-dependent Src activation.

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Biosynthesis of the insulin receptor in rat adipose cells. Intracellular processing of the Mr-190 000 pro-receptor

Biochemical Journal ◽

10.1042/bj2320071 ◽

1985 ◽

Vol 232 (1) ◽

pp. 71-78 ◽

Cited By ~ 15

Author(s):

J A Hedo ◽

I A Simpson

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Insulin Receptor ◽

Golgi Complex ◽

Microsomal Fraction ◽

Primary Cultures ◽

Sodium Dodecyl ◽

High Density ◽

Low Density ◽

Adipose Cells

We investigated the biosynthesis of the insulin receptor in primary cultures of isolated rat adipose cells. Cells were pulse-chase-labelled with [3H]mannose, and at intervals samples were homogenized. Three subcellular membrane fractions were prepared by differential centrifugation: high-density microsomal (endoplasmic-reticulum-enriched), low-density microsomal (Golgi-enriched), and plasma membranes. After detergent solubilization, the insulin receptors were immunoprecipitated with anti-receptor antibodies and analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and autoradiography. After a 30 min pulse-label [3H]mannose first appeared in a band of Mr 190 000. More than 80% of the Mr-190 000 component was recovered in the microsomal fractions. Its intensity reached a maximum at 1 h in the high-density microsomal fraction and at 2 h in the low-density microsomal fraction, and thereafter declined rapidly (t 1/2 approx. 3 h) in both fractions. In the plasma-membrane fraction, the radioactivity in the major receptor subunits, of Mr 135 000 (alpha) and 95 000 (beta), rose steadily during the chase and reached a maximum at 6 h. The Mr-190 000 precursor could also be detected in the high-density microsomal fraction by affinity cross-linking to 125I-insulin. In the presence of monensin, a cationic ionophore that interferes with intracellular transport within the Golgi complex, the processing of the Mr-190 000 precursor into the alpha and beta subunits was completely inhibited. Our results suggest that the Mr-190 000 pro-receptor originates in the endoplasmic reticulum and is subsequently transferred to the Golgi complex. Maturation of the pro-receptor does not seem to be necessary for the expression of the insulin-binding site. Processing of the precursor into the mature receptor subunits appears to occur during the transfer of the pro-receptor from the Golgi complex to the plasma membrane.

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RAB6 and microtubules restrict protein secretion to focal adhesions

The Journal of Cell Biology ◽

10.1083/jcb.201805002 ◽

2019 ◽

Vol 218 (7) ◽

pp. 2215-2231 ◽

Cited By ~ 32

Author(s):

Lou Fourriere ◽

Amal Kasri ◽

Nelly Gareil ◽

Sabine Bardin ◽

Hugo Bousquet ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Cell Surface ◽

Protein Secretion ◽

Golgi Complex ◽

Essential Role ◽

Focal Adhesions ◽

Secreted Proteins ◽

Cell Compartments ◽

Secretion Assay

To ensure their homeostasis and sustain differentiated functions, cells continuously transport diverse cargos to various cell compartments and in particular to the cell surface. Secreted proteins are transported along intracellular routes from the endoplasmic reticulum through the Golgi complex before reaching the plasma membrane along microtubule tracks. Using a synchronized secretion assay, we report here that exocytosis does not occur randomly at the cell surface but on localized hotspots juxtaposed to focal adhesions. Although microtubules are involved, the RAB6-dependent machinery plays an essential role. We observed that, irrespective of the transported cargos, most post-Golgi carriers are positive for RAB6 and that its inactivation leads to a broad reduction of protein secretion. RAB6 may thus be a general regulator of post-Golgi secretion.

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Kv2 potassium channels form endoplasmic reticulum/plasma membrane junctions via interaction with VAPA and VAPB

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1805757115 ◽

2018 ◽

Vol 115 (31) ◽

pp. E7331-E7340 ◽

Cited By ~ 46

Author(s):

Ben Johnson ◽

Ashley N. Leek ◽

Laura Solé ◽

Emily E. Maverick ◽

Tim P. Levine ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Membrane Trafficking ◽

Neuronal Cultures ◽

Physical Relationship ◽

Membrane Contact Sites ◽

Contact Sites ◽

Hek Cells ◽

C Terminus ◽

Membrane Contact

Kv2.1 exhibits two distinct forms of localization patterns on the neuronal plasma membrane: One population is freely diffusive and regulates electrical activity via voltage-dependent K+ conductance while a second one localizes to micrometer-sized clusters that contain densely packed, but nonconducting, channels. We have previously established that these clusters represent endoplasmic reticulum/plasma membrane (ER/PM) junctions that function as membrane trafficking hubs and that Kv2.1 plays a structural role in forming these membrane contact sites in both primary neuronal cultures and transfected HEK cells. Clustering and the formation of ER/PM contacts are regulated by phosphorylation within the channel C terminus, offering cells fast, dynamic control over the physical relationship between the cortical ER and PM. The present study addresses the mechanisms by which Kv2.1 and the related Kv2.2 channel interact with the ER membrane. Using proximity-based biotinylation techniques in transfected HEK cells we identified ER VAMP-associated proteins (VAPs) as potential Kv2.1 interactors. Confirmation that Kv2.1 and -2.2 bind VAPA and VAPB employed colocalization/redistribution, siRNA knockdown, and Förster resonance energy transfer (FRET)-based assays. CD4 chimeras containing sequence from the Kv2.1 C terminus were used to identify a noncanonical VAP-binding motif. VAPs were first identified as proteins required for neurotransmitter release in Aplysia and are now known to be abundant scaffolding proteins involved in membrane contact site formation throughout the ER. The VAP interactome includes AKAPs, kinases, membrane trafficking machinery, and proteins regulating nonvesicular lipid transport from the ER to the PM. Therefore, the Kv2-induced VAP concentration at ER/PM contact sites is predicted to have wide-ranging effects on neuronal cell biology.

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Segregation in the Golgi complex precedes export of endolysosomal proteins in distinct transport carriers

The Journal of Cell Biology ◽

10.1083/jcb.201707172 ◽

2017 ◽

Vol 216 (12) ◽

pp. 4141-4151 ◽

Cited By ~ 38

Author(s):

Yu Chen ◽

David C. Gershlick ◽

Sang Yoon Park ◽

Juan S. Bonifacino

Keyword(s):

Plasma Membrane ◽

Membrane Proteins ◽

Golgi Complex ◽

Adaptor Proteins ◽

Transmembrane Domains ◽

The Other ◽

Present Evidence ◽

Sorting Signals ◽

Vesicular Carriers ◽

Classical Models

Biosynthetic sorting of newly synthesized transmembrane cargos to endosomes and lysosomes is thought to occur at the TGN through recognition of sorting signals in the cytosolic tails of the cargos by adaptor proteins, leading to cargo packaging into coated vesicles destined for the endolysosomal system. Here we present evidence for a different mechanism in which two sets of endolysosomal proteins undergo early segregation to distinct domains of the Golgi complex by virtue of the proteins’ luminal and transmembrane domains. Proteins in one Golgi domain exit into predominantly vesicular carriers by interaction of sorting signals with adaptor proteins, but proteins in the other domain exit into predominantly tubular carriers shared with plasma membrane proteins, independently of signal–adaptor interactions. These findings demonstrate that sorting of endolysosomal proteins begins at an earlier stage and involves mechanisms that partly differ from those described by classical models.

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