The recycling pathway of protein ERGIC-53 and dynamics of the ER-Golgi intermediate compartment

1998 ◽  
Vol 111 (22) ◽  
pp. 3411-3425 ◽  
Author(s):  
J. Klumperman ◽  
A. Schweizer ◽  
H. Clausen ◽  
B.L. Tang ◽  
W. Hong ◽  
...  
Keyword(s):  
Golgi Apparatus ◽  
Secretory Pathway ◽  
Immunofluorescence Microscopy ◽  
Gradient Centrifugation ◽  
Membrane System ◽  
Cell Periphery ◽  
Dynamic Membrane ◽  
Early Secretory Pathway ◽  
Intermediate Compartment ◽  
Recycling Pathway

To establish recycling routes in the early secretory pathway we have studied the recycling of the ER-Golgi intermediate compartment (ERGIC) marker ERGIC-53 in HepG2 cells. Immunofluorescence microscopy showed progressive concentration of ERGIC-53 in the Golgi area at 15 degreesC. Upon rewarming to 37 degreesC ERGIC-53 redistributed into the cell periphery often via tubular processes that largely excluded anterograde transported albumin. Immunogold labeling of cells cultured at 37 degreesC revealed ERGIC-53 predominantly in characteristic beta-COP-positive tubulo-vesicular clusters both near the Golgi apparatus and in the cell periphery. Concentration of ERGIC-53 at 15 degreesC resulted from both accumulation of ERGIC-53 in the ERGIC and movement of ERGIC membranes closer to the Golgi apparatus. Upon rewarming to 37 degreesC the labeling of ERGIC-53 in the ERGIC rapidly returned to normal levels whereas ERGIC-53's labeling in the cis-Golgi was unchanged. Temperature manipulations had no effect on the average number of ERGIC-53 clusters. Density gradient centrifugation indicated that the surplus ERGIC-53 accumulating in the ERGIC at 15 degreesC was rapidly transported to the ER upon rewarming. These results suggest that the ERGIC is a dynamic membrane system composed of a constant average number of clusters and that the major recycling pathway of ERGIC-53 bypasses the Golgi apparatus.

scholarly journals Branched Actin Maintains Acetylated Microtubule Network in the Early Secretory Pathway

Cells ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 15
Author(s):  
Azumi Yoshimura ◽  
Stéphanie Miserey-Lenkei ◽  
Evelyne Coudrier ◽  
Bruno Goud
Keyword(s):  
Golgi Apparatus ◽  
Transport Process ◽  
Actin Polymerization ◽  
Secretory Pathway ◽  
Microtubule Network ◽  
Early Secretory Pathway ◽  
Golgi Transport ◽  
Er Exit Sites ◽  
Intermediate Compartment ◽  
Stable Network

In the early secretory pathway, the delivery of anterograde cargoes from the endoplasmic reticulum (ER) exit sites (ERES) to the Golgi apparatus is a multi-step transport process occurring via the ER-Golgi intermediate compartment (IC, also called ERGIC). While the role microtubules in ER-to-Golgi transport has been well established, how the actin cytoskeleton contributes to this process remains poorly understood. Here, we report that Arp2/3 inhibition affects the network of acetylated microtubules around the Golgi and induces the accumulation of unusually long RAB1/GM130-positive carriers around the centrosome. These long carriers are less prone to reach the Golgi apparatus, and arrival of anterograde cargoes to the Golgi is decreased upon Arp2/3 inhibition. Our data suggest that Arp2/3-dependent actin polymerization maintains a stable network of acetylated microtubules, which ensures efficient cargo trafficking at the late stage of ER to Golgi transport.

scholarly journals Processing of prothrombin in the secretory pathway

1991 ◽  
Vol 277 (1) ◽  
pp. 59-65 ◽  
Author(s):  
C Stanton ◽  
R Taylor ◽  
R Wallin
Keyword(s):  
Golgi Apparatus ◽  
Secretory Pathway ◽  
Gradient Centrifugation ◽  
Serine Proteinase ◽  
Sucrose Density Gradient ◽  
Sucrose Density Gradient Centrifugation ◽  
Carboxylase Activity ◽  
Parent Molecule ◽  
Early Processing ◽  
In Cis

Antibodies raised against plasma prothrombin and the prothrombin propeptide were used to identify prothrombin precursors in rough and smooth microsomes and in the Golgi apparatus. The data demonstrate that the propeptide is part of the prothrombin molecule when undergoing a variety of modifications in the Golgi apparatus. It is shown that these modifications result in an increase in the apparent molecular mass of the prothrombin precursor from 78 kDa in early processing to 83 kDa in late processing. The 83 kDa prothrombin precursor was not recognized by the anti-propeptide antiserum and most likely represents the final product of the precursor in the secretory pathway. Evidence is presented that the propeptide is released from the parent molecule in the Golgi apparatus by a membrane-bound Ca(2+)-dependent serine proteinase(s) with characteristics similar to those of the proalbumin-to-albumin-converting enzyme. Vitamin K-dependent carboxylase activity was measured in membrane fragments obtained from the Golgi apparatus preparation. Sucrose-density-gradient centrifugation and the use of marker enzymes showed that carboxylase activity was highest in fractions enriched in cis-Golgi cisternae. Two different synthetic peptides were used as substrates for the carboxylase. These peptides were from the N-terminal and the C-terminal part of the gamma-carboxyglutamic acid (Gla) region of prothrombin. It is shown that the N-terminal and the C-terminal peptides were preferred as substrates for the carboxylase in the microsomal and the Golgi apparatus preparations respectively. It is also shown that the prothrombin precursor acquires negative charges in the Golgi apparatus that do not result from addition of sugars in late processing. These negative charges could be eliminated by thermal decarboxylation, suggesting that Gla residues may also be synthesized in late processing.

scholarly journals MIA3/TANGO1 enables efficient secretion by constraining COPII vesicle budding

2021 ◽  
Author(s):  
Janine McCaughey ◽  
Judith M. Mantell ◽  
Chris R. Neal ◽  
Kate Heesom ◽  
David J. Stephens
Keyword(s):  
Endoplasmic Reticulum ◽  
Light Microscopy ◽  
Secretory Pathway ◽  
Genome Engineering ◽  
High Volume ◽  
Vesicle Budding ◽  
Early Secretory Pathway ◽  
Copii Vesicle ◽  
Intermediate Compartment ◽  
Golgi Organization

AbstractComplex machinery is required to drive secretory cargo export from the endoplasmic reticulum. In vertebrates, this includes transport and Golgi organization protein 1 (TANGO1), encoded by the Mia3 gene. Here, using genome engineering of human cells light microscopy, secretion assays, and proteomics, we show loss of Mia3/TANGO1 results in formation of numerous vesicles and a loss of early secretory pathway integrity. This restricts secretion not only of large proteins like procollagens but of all types of secretory cargo. Our data shows that Mia3/TANGO1 constrains the propensity of COPII to form vesicles promoting instead the formation of the ER-Golgi intermediate compartment. Thus, Mia3/TANGO1 facilities the secretion of complex and high volume cargoes from vertebrate cells.

scholarly journals Poliovirus Infection Transiently Increases COPII Vesicle Budding

2012 ◽  
Vol 86 (18) ◽  
pp. 9675-9682 ◽  
Author(s):  
Meg Trahey ◽  
Hyung Suk Oh ◽  
Craig E. Cameron ◽  
Jesse C. Hay
Keyword(s):  
Golgi Apparatus ◽  
Secretory Pathway ◽  
Vesicle Formation ◽  
Vesicle Budding ◽  
Precursor Pool ◽  
Copii Vesicle ◽  
Early Increase ◽  
Vesicle Biogenesis ◽  
Intermediate Compartment ◽  
Copii Vesicles

Poliovirus (PV) requires membranes of the host cell's secretory pathway to generate replication complexes (RCs) for viral RNA synthesis. Recent work identified the intermediate compartment and the Golgi apparatus as the precursors of the replication “organelles” of PV (N. Y. Hsu et al., Cell 141:799–811, 2010). In this study, we examined the effect of PV on COPII vesicles, the secretory cargo carriers that bud from the endoplasmic reticulum and homotypically fuse to form the intermediate compartment that matures into the Golgi apparatus. We found that infection by PV results in a biphasic change in functional COPII vesicle biogenesis in cells, with an early enhancement and subsequent inhibition. Concomitant with the early increase in COPII vesicle formation, we found an increase in the membrane fraction of Sec16A, a key regulator of COPII vesicle formation. We suggest that the early burst in COPII vesicle formation detected benefits PV by increasing the precursor pool required for the formation of its RCs.

scholarly journals The Function of the Intermediate Compartment in Pre-Golgi Trafficking Involves its Stable Connection with the Centrosome

2009 ◽  
Vol 20 (20) ◽  
pp. 4458-4470 ◽  
Author(s):  
Michaël Marie ◽  
Hege A. Dale ◽  
Ragna Sannerud ◽  
Jaakko Saraste
Keyword(s):  
Secretory Pathway ◽  
Focal Point ◽  
Brefeldin A ◽  
Temperature Sensitive ◽  
Cell Periphery ◽  
Functional Connection ◽  
Centrosome Separation ◽  
Membrane Compartment ◽  
Intermediate Compartment ◽  
Endosomal System

Because the functional borders of the intermediate compartment (IC) are not well defined, the spatial map of the transport machineries operating between the endoplasmic reticulum (ER) and the Golgi apparatus remains incomplete. Our previous studies showed that the IC consists of interconnected vacuolar and tubular parts with specific roles in pre-Golgi trafficking. Here, using live cell imaging, we demonstrate that the tubules containing the GTPase Rab1A create a long-lived membrane compartment around the centrosome. Separation of this pericentrosomal domain of the IC from the Golgi ribbon, due to centrosome motility, revealed that it contains a distinct pool of COPI coats and acts as a temperature-sensitive way station in post-ER trafficking. However, unlike the Golgi, the pericentrosomal IC resists the disassembly of COPI coats by brefeldin A, maintaining its juxtaposition with the endocytic recycling compartment, and operation as the focal point of a dynamic tubular network that extends to the cell periphery. These results provide novel insight into the compartmental organization of the secretory pathway and Golgi biogenesis. Moreover, they reveal a direct functional connection between the IC and the endosomal system, which evidently contributes to unconventional transport of the cystic fibrosis transmembrane conductance regulator to the cell surface.

scholarly journals The Cargo Receptors Surf4, Endoplasmic Reticulum-Golgi Intermediate Compartment (ERGIC)-53, and p25 Are Required to Maintain the Architecture of ERGIC and Golgi

2008 ◽  
Vol 19 (5) ◽  
pp. 1976-1990 ◽  
Author(s):  
Sandra Mitrovic ◽  
Houchaima Ben-Tekaya ◽  
Eva Koegler ◽  
Jean Gruenberg ◽  
Hans-Peter Hauri
Keyword(s):  
Endoplasmic Reticulum ◽  
Secretory Pathway ◽  
Brefeldin A ◽  
Matrix Proteins ◽  
Anterograde Transport ◽  
Early Secretory Pathway ◽  
Partial Redistribution ◽  
Intermediate Compartment ◽  
Human Orthologue ◽  
Golgi Matrix

Rapidly cycling proteins of the early secretory pathway can operate as cargo receptors. Known cargo receptors are abundant proteins, but it remains mysterious why their inactivation leads to rather limited secretion phenotypes. Studies of Surf4, the human orthologue of the yeast cargo receptor Erv29p, now reveal a novel function of cargo receptors. Surf4 was found to interact with endoplasmic reticulum-Golgi intermediate compartment (ERGIC)-53 and p24 proteins. Silencing Surf4 together with ERGIC-53 or silencing the p24 family member p25 induced an identical phenotype characterized by a reduced number of ERGIC clusters and fragmentation of the Golgi apparatus without effect on anterograde transport. Live imaging showed decreased stability of ERGIC clusters after knockdown of p25. Silencing of Surf4/ERGIC-53 or p25 resulted in partial redistribution of coat protein (COP) I but not Golgi matrix proteins to the cytosol and partial resistance of the cis-Golgi to brefeldin A. These findings imply that cargo receptors are essential for maintaining the architecture of ERGIC and Golgi by controlling COP I recruitment.

scholarly journals The small GTP-binding protein Rho1p is localized on the Golgi apparatus and post-Golgi vesicles in Saccharomyces cerevisiae.

1991 ◽  
Vol 115 (2) ◽  
pp. 309-319 ◽  
Author(s):  
M McCaffrey ◽  
J S Johnson ◽  
B Goud ◽  
A M Myers ◽  
J Rossier ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Golgi Apparatus ◽  
Immunofluorescence Microscopy ◽  
Cell Periphery ◽  
Related Protein ◽  
Wild Type ◽  
Golgi Vesicles ◽  
Golgi Transport ◽  
Transport Vesicles ◽  
Punctate Pattern

In Saccharomyces cerevisiae the ras-related protein Rho1p is essentially the only target for ADP-ribosylation by exoenzyme C3 of Clostridium botulinum. Using C3 to detect Rho1p in subcellular fractions, Rho1p was found primarily in the 10,000 g pellet (P2) containing large organelles; small amounts also were detected in the 100,000 g pellet (P3), and cytosol. When P2 organelles were separated in sucrose density gradients Rho1p comigrated with the Kex-2 activity, a late Golgi marker. Rho1p distribution was shifted from P2 to P3 in several mutants that accumulate post-Golgi vesicles. Rho1p comigrated with post-Golgi transport vesicles during fractionation of P3 organelles from wild-type or sec6 cells. Vesicles containing Rho1p were of the same size but different density than those bearing Sec4p, a ras-related protein located both on post-Golgi vesicles and the plasma membrane. Immunofluorescence microscopy detected Rho1p as a punctate pattern, with signal concentrated towards the cell periphery and in the bud. Thus, in S. cerevisiae Rho1p resides primarily in the Golgi apparatus, and also in vesicles that are likely to be early post-Golgi vesicles.

scholarly journals p53/58 Binds COPI and Is Required for Selective Transport through the Early Secretory Pathway

1997 ◽  
Vol 137 (3) ◽  
pp. 581-593 ◽  
Author(s):  
Ellen J. Tisdale ◽  
Helen Plutner ◽  
Jeanne Matteson ◽  
William E. Balch
Keyword(s):  
Secretory Pathway ◽  
Transmembrane Protein ◽  
Cytoplasmic Tail ◽  
Cell Periphery ◽  
Vesicular Stomatitis Virus Glycoprotein ◽  
Anterograde Transport ◽  
Selective Transport ◽  
Early Secretory Pathway ◽  
Vesicular Stomatitis ◽  
Retrograde Traffic

p53/58 is a transmembrane protein that continuously recycles between the ER and pre-Golgi intermediates composed of vesicular-tubular clusters (VTCs) found in the cell periphery and at the cis face of the Golgi complex. We have generated an antibody that uniquely recognizes the p53/58 cytoplasmic tail. Here we present evidence that this antibody arrests the anterograde transport of vesicular stomatitis virus glycoprotein and leads to the accumulation of p58 in preGolgi intermediates. Consistent with a role for the KKXX retrieval motif found at the cytoplasmic carboxyl terminus of p53/58 in retrograde traffic, inhibition of transport through VTCs correlates with the ability of the antibody to block recruitment of COPI coats to the p53/58 cytoplasmic tail and to p53/58-containing membranes. We suggest that p53/58 function may be required for the coupled exchange of COPII for COPI coats during segregation of anterograde and retrograde transported proteins.

scholarly journals The transmembrane protein p23 contributes to the organization of the Golgi apparatus

2000 ◽  
Vol 113 (6) ◽  
pp. 1043-1057 ◽  
Author(s):  
M. Rojo ◽  
G. Emery ◽  
V. Marjomaki ◽  
A.W. McDowall ◽  
R.G. Parton ◽  
...  
Keyword(s):  
Golgi Apparatus ◽  
Secretory Pathway ◽  
Transmembrane Protein ◽  
Ectopic Expression ◽  
Retrograde Transport ◽  
Early Secretory Pathway ◽  
Golgi Membranes ◽  
Constant Diameter ◽  
Golgi Network

In previous studies we have shown that p23, a member of the p24-family of small transmembrane proteins, is highly abundant in membranes of the cis-Golgi network (CGN), and is involved in sorting/trafficking in the early secretory pathway. In the present study, we have further investigated the role of p23 after ectopic expression. We found that ectopically expressed p23 folded and oligomerized properly, even after overexpression. However, in contrast to endogenous p23, exogenous p23 molecules did not localize to the CGN, but induced a significant expansion of characteristic smooth ER membranes, where they accumulated in high amounts. This ER-derived, p23-rich subdomain displayed a highly regular morphology, consisting of tubules and/or cisternae of constant diameter, which were reminiscent of the CGN membranes containing p23 in control cells. The expression of exogenous p23 also led to the specific relocalization of endogenous p23, but not of other proteins, to these specialized ER-derived membranes. Relocalization of p23 modified the ultrastructure of the CGN and Golgi membranes, but did not affect anterograde and retrograde transport reactions to any significant extent. We conclude (i) that p23 has a morphogenic activity that contributes to the morphology of CGN-membranes; and (ii) that the presence of p23 in the CGN is necessary for the proper organization of the Golgi apparatus.

scholarly journals Assembly and Cellular Exit of Coronaviruses: Hijacking an Unconventional Secretory Pathway from the Pre-Golgi Intermediate Compartment via the Golgi Ribbon to the Extracellular Space

Cells ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 503
Author(s):  
Jaakko Saraste ◽  
Kristian Prydz
Keyword(s):  
Endoplasmic Reticulum ◽  
Extracellular Space ◽  
Secretory Pathway ◽  
Secretory Process ◽  
Cell Periphery ◽  
Direct Connection ◽  
Recycling Endosomes ◽  
Functional Connections ◽  
Intermediate Compartment ◽  
Golgi Ribbon

Coronaviruses (CoVs) assemble by budding into the lumen of the intermediate compartment (IC) at the endoplasmic reticulum (ER)-Golgi interface. However, why CoVs have chosen the IC as their intracellular site of assembly and how progeny viruses are delivered from this compartment to the extracellular space has remained unclear. Here we address these enigmatic late events of the CoV life cycle in light of recently described properties of the IC. Of particular interest are the emerging spatial and functional connections between IC elements and recycling endosomes (REs), defined by the GTPases Rab1 and Rab11, respectively. The establishment of IC-RE links at the cell periphery, around the centrosome and evidently also at the noncompact zones of the Golgi ribbon indicates that—besides traditional ER-Golgi communication—the IC also promotes a secretory process that bypasses the Golgi stacks, but involves its direct connection with the endocytic recycling system. The initial confinement of CoVs to the lumen of IC-derived large transport carriers and their preferential absence from Golgi stacks is consistent with the idea that they exit cells following such an unconventional route. In fact, CoVs may share this pathway with other intracellularly budding viruses, lipoproteins, procollagen, and/or protein aggregates experimentally introduced into the IC lumen.

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