scholarly journals cis-Golgi proteins accumulate near the ER exit sites and act as the scaffold for Golgi regeneration after brefeldin A treatment in tobacco BY-2 cells

2012 ◽  
Vol 23 (16) ◽  
pp. 3203-3214 ◽  
Author(s):  
Yoko Ito ◽  
Tomohiro Uemura ◽  
Keiko Shoda ◽  
Masaru Fujimoto ◽  
Takashi Ueda ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Fluorescent Proteins ◽  
Plant Cells ◽  
Brefeldin A ◽  
Eukaryotic Cells ◽  
Er Exit Sites ◽  
Intermediate Compartment ◽  
Golgi Protein ◽  
And Function

The Golgi apparatus forms stacks of cisternae in many eukaryotic cells. However, little is known about how such a stacked structure is formed and maintained. To address this question, plant cells provide a system suitable for live-imaging approaches because individual Golgi stacks are well separated in the cytoplasm. We established tobacco BY-2 cell lines expressing multiple Golgi markers tagged by different fluorescent proteins and observed their responses to brefeldin A (BFA) treatment and BFA removal. BFA treatment disrupted cis, medial, and trans cisternae but caused distinct relocalization patterns depending on the proteins examined. Medial- and trans-Golgi proteins, as well as one cis-Golgi protein, were absorbed into the endoplasmic reticulum (ER), but two other cis-Golgi proteins formed small punctate structures. After BFA removal, these puncta coalesced first, and then the Golgi stacks regenerated from them in the cis-to-trans order. We suggest that these structures have a property similar to the ER-Golgi intermediate compartment and function as the scaffold of Golgi regeneration.

Dynamic association of cytoplasmic dynein heavy chain 1a with the Golgi apparatus and intermediate compartment

1999 ◽  
Vol 112 (24) ◽  
pp. 4673-4685 ◽  
Author(s):  
C. Roghi ◽  
V.J. Allan
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Heavy Chain ◽  
Brefeldin A ◽  
Cytoplasmic Dynein ◽  
Intracellular Location ◽  
Dynein Heavy Chain ◽  
Intermediate Compartment ◽  
And Function ◽  
Dynactin Complex

Microtubule motors, such as the minus end-directed motor, cytoplasmic dynein, play an important role in maintaining the integrity, intracellular location, and function of the Golgi apparatus, as well as in the translocation of membrane between the endoplasmic reticulum and Golgi apparatus. We have immunolocalised conventional cytoplasmic dynein heavy chain to the Golgi apparatus in cultured vertebrate cells. In addition, we present evidence that cytoplasmic dynein heavy chain cycles constitutively between the endoplasmic reticulum and Golgi apparatus: it colocalises partially with the intermediate compartment, it is found on nocodazole-induced peripheral Golgi elements and, most strikingly, on Brefeldin A-induced tubules that are moving towards microtubule plus ends. The direction of movement of membrane between the endoplasmic reticulum and Golgi apparatus is therefore unlikely to be regulated by controlling motor-membrane interactions: rather, the motors probably remain bound throughout the whole cycle, with their activity being modulated instead. We also report that the overexpression of p50/dynamitin results in the loss of cytoplasmic dynein heavy chain from the membrane of peripheral Golgi elements. These results explain how dynamitin overexpression causes the inhibition of endoplasmic reticulum-to-Golgi transport complex movement towards the centrosomal region, and support the general model that an intact dynactin complex is required for cytoplasmic dynein binding to all cargoes.

Brefeldin A-induced disassembly of the Golgi apparatus is followed by disruption of the endoplasmic reticulum in plant cells

1994 ◽  
Vol 45 (10) ◽  
pp. 1347-1351 ◽  
Author(s):  
Janey Henderson ◽  
Béatrice Stiat-Jeunemaitre ◽  
Richard Napier ◽  
Chris Hawes
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Plant Cells ◽  
Brefeldin A

scholarly journals Maintenance of Golgi structure and function depends on the integrity of ER export

2001 ◽  
Vol 155 (4) ◽  
pp. 557-570 ◽  
Author(s):  
Theresa H. Ward ◽  
Roman S. Polishchuk ◽  
Steve Caplan ◽  
Koret Hirschberg ◽  
Jennifer Lippincott-Schwartz
Keyword(s):  
Steady State ◽  
Golgi Apparatus ◽  
Membrane Trafficking ◽  
Imaging Techniques ◽  
Brefeldin A ◽  
Er Exit Sites ◽  
Selective Membrane ◽  
Golgi Structure ◽  
And Function ◽  
Er Export

The Golgi apparatus comprises an enormous array of components that generate its unique architecture and function within cells. Here, we use quantitative fluorescence imaging techniques and ultrastructural analysis to address whether the Golgi apparatus is a steady-state or a stable organelle. We found that all classes of Golgi components are dynamically associated with this organelle, contrary to the prediction of the stable organelle model. Enzymes and recycling components are continuously exiting and reentering the Golgi apparatus by membrane trafficking pathways to and from the ER, whereas Golgi matrix proteins and coatomer undergo constant, rapid exchange between membrane and cytoplasm. When ER to Golgi transport is inhibited without disrupting COPII-dependent ER export machinery (by brefeldin A treatment or expression of Arf1[T31N]), the Golgi structure disassembles, leaving no residual Golgi membranes. Rather, all Golgi components redistribute into the ER, the cytoplasm, or to ER exit sites still active for recruitment of selective membrane-bound and peripherally associated cargos. A similar phenomenon is induced by the constitutively active Sar1[H79G] mutant, which has the additional effect of causing COPII-associated membranes to cluster to a juxtanuclear region. In cells expressing Sar1[T39N], a constitutively inactive form of Sar1 that completely disrupts ER exit sites, Golgi glycosylation enzymes, matrix, and itinerant proteins all redistribute to the ER. These results argue against the hypothesis that the Golgi apparatus contains stable components that can serve as a template for its biogenesis. Instead, they suggest that the Golgi complex is a dynamic, steady-state system, whose membranes can be nucleated and are maintained by the activities of the Sar1–COPII and Arf1–coatomer systems.

scholarly journals GS15 Forms a SNARE Complex with Syntaxin 5, GS28, and Ykt6 and Is Implicated in Traffic in the Early Cisternae of the Golgi Apparatus

2002 ◽  
Vol 13 (10) ◽  
pp. 3493-3507 ◽  
Author(s):  
Yue Xu ◽  
Sally Martin ◽  
David E. James ◽  
Wanjin Hong
Keyword(s):  
Golgi Apparatus ◽  
Brefeldin A ◽  
Snare Complex ◽  
Er Exit Sites ◽  
And Function ◽  
Mutant Forms ◽  
Interfering Rna ◽  
Kdel Receptor ◽  
Golgi Matrix

The subcellular localization, interacting partners, and function of GS15, a Golgi SNARE, remain to be established. In our present study, it is revealed that unlike proteins (Bet1 and the KDEL receptor) cycling between the Golgi and the intermediate compartment (IC, inclusive of the ER exit sites), GS15 is not redistributed into the IC upon incubation at 15°C or when cells are treated with brefeldin A. Immuno-electron microscopy (immuno-EM) reveals that GS15 is mainly found in the medial-cisternae of the Golgi apparatus and adjacent tubulo-vesicular elements. Coimmunoprecipitation experiments suggest that GS15 exists in a distinct SNARE complex that contains SNAREs (syntaxin5, GS28, and Ykt6) that are implicated in both ER-to-Golgi and intra-Golgi transport but not with SNAREs involved exclusively in ER-to-Golgi traffic. Furthermore, components of COPI coat can be selectively coimmunoprecipitated with GS15 from Golgi extracts. Overexpression of mutant forms of GS15 affects the normal distribution of cis- and medial-Golgi proteins (GS28, syntaxin 5, and Golgi mannosidase II), whereas proteins of the trans-Golgi and TGN (Vti1-rp2/Vti1a and syntaxin 6) and Golgi matrix/scaffold (GM130 and p115) are less affected. When the level of GS15 is reduced by duplex 21-nt small interfering RNA (siRNA)-mediated knockdown approach, diverse markers of the Golgi apparatus are redistributed into small dotty and diffuse labeling, suggesting an essential role of GS15 in the Golgi apparatus.

scholarly journals Functional Symmetry of Endomembranes

2007 ◽  
Vol 18 (4) ◽  
pp. 1430-1436 ◽  
Author(s):  
Jaakko Saraste ◽  
Bruno Goud
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Cell Activation ◽  
Membrane System ◽  
Eukaryotic Cells ◽  
Membrane Recycling ◽  
Major Task ◽  
Late Endosomes ◽  
Intermediate Compartment ◽  
Functional Symmetry

In higher eukaryotic cells pleiomorphic compartments composed of vacuoles, tubules and vesicles move from the endoplasmic reticulum (ER) and the plasma membrane to the cell center, operating in early biosynthetic trafficking and endocytosis, respectively. Besides transporting cargo to the Golgi apparatus and lysosomes, a major task of these compartments is to promote extensive membrane recycling. The endocytic membrane system is traditionally divided into early (sorting) endosomes, late endosomes and the endocytic recycling compartment (ERC). Recent studies on the intermediate compartment (IC) between the ER and the Golgi apparatus suggest that it also consists of peripheral (“early”) and centralized (“late”) structures, as well as a third component, designated here as the biosynthetic recycling compartment (BRC). We propose that the ERC and the BRC exist as long-lived “mirror compartments” at the cell center that also share the ability to expand and become mobilized during cell activation. These considerations emphasize the functional symmetry of endomembrane compartments, which provides a basis for the membrane rearrangements taking place during cell division, polarization, and differentiation.

scholarly journals Capacity of the Golgi Apparatus for Biogenesis from the Endoplasmic Reticulum

2003 ◽  
Vol 14 (12) ◽  
pp. 5011-5018 ◽  
Author(s):  
Sapna Puri ◽  
Adam D. Linstedt
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Self Assembly ◽  
De Novo ◽  
Brefeldin A ◽  
The Other ◽  
Matrix Proteins ◽  
Er Export ◽  
Golgi Matrix

It is unclear whether the mammalian Golgi apparatus can form de novo from the ER or whether it requires a preassembled Golgi matrix. As a test, we assayed Golgi reassembly after forced redistribution of Golgi matrix proteins into the ER. Two conditions were used. In one, ER redistribution was achieved using a combination of brefeldin A (BFA) to cause Golgi collapse and H89 to block ER export. Unlike brefeldin A alone, which leaves matrix proteins in relatively large remnant structures outside the ER, the addition of H89 to BFA-treated cells caused ER accumulation of all Golgi markers tested. In the other, clofibrate treatment induced ER redistribution of matrix and nonmatrix proteins. Significantly, Golgi reassembly after either treatment was robust, implying that the Golgi has the capacity to form de novo from the ER. Furthermore, matrix proteins reemerged from the ER with faster ER exit rates. This, together with the sensitivity of BFA remnants to ER export blockade, suggests that presence of matrix proteins in BFA remnants is due to cycling via the ER and preferential ER export rather than their stable assembly in a matrix outside the ER. In summary, the Golgi apparatus appears capable of efficient self-assembly.

scholarly journals Molecular cloning, characterization, subcellular localization and dynamics of p23, the mammalian KDEL receptor.

1993 ◽  
Vol 120 (2) ◽  
pp. 325-338 ◽  
Author(s):  
B L Tang ◽  
S H Wong ◽  
X L Qi ◽  
S H Low ◽  
W Hong
Keyword(s):  
Golgi Apparatus ◽  
Brefeldin A ◽  
Cell Types ◽  
In Vitro Translation ◽  
Tubular Structures ◽  
Human Sequence ◽  
Intermediate Compartment ◽  
Membrane Spanning ◽  
Membrane Spanning Domains

We have isolated a cDNA clone (mERD2) for the mammalian (bovine) homologue of the yeast ERD2 gene, which codes for the yeast HDEL receptor. The deduced amino acid sequence bears extensive homology to its yeast counterpart and is almost identical to a previously described human sequence. The sequence predicts a very hydrophobic protein with multiple membrane spanning domains, as confirmed by analysis of the in vitro translation product. The protein encoded by mERD2 (p23) has widespread occurrence, being present in all the cell types examined. p23 was localized to the cis-side of the Golgi apparatus and to a spotty intermediate compartment which mediates ER to Golgi transport. A majority of the intracellular staining could be accumulated in the intermediate compartment by a low temperature (15 degrees C) or brefeldin A. During recovery from these treatments, the spotty intermediate compartment staining of p23 was shifted to the perinuclear staining of the Golgi apparatus and tubular structures marked by p23 were observed. These tubular structures may serve to mediate transport between the intermediate compartment and the Golgi apparatus.

scholarly journals The discrepancy between presenilin subcellular localization and γ-secretase processing of amyloid precursor protein

2001 ◽  
Vol 154 (4) ◽  
pp. 731-740 ◽  
Author(s):  
Philippe Cupers ◽  
Mustapha Bentahir ◽  
Katleen Craessaerts ◽  
Isabelle Orlans ◽  
Hugo Vanderstichele ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Surface ◽  
Amyloid Precursor Protein ◽  
Primary Cultures ◽  
Brefeldin A ◽  
Precursor Protein ◽  
Subcellular Compartment ◽  
Marker Proteins ◽  
Intermediate Compartment ◽  
The Relationship

We investigated the relationship between PS1 and γ-secretase processing of amyloid precursor protein (APP) in primary cultures of neurons. Increasing the amount of APP at the cell surface or towards endosomes did not significantly affect PS1-dependent γ-secretase cleavage, although little PS1 is present in those subcellular compartments. In contrast, almost no γ-secretase processing was observed when holo-APP or APP-C99, a direct substrate for γ-secretase, were specifically retained in the endoplasmic reticulum (ER) by a double lysine retention motif. Nevertheless, APP-C99-dilysine (KK) colocalized with PS1 in the ER. In contrast, APP-C99 did not colocalize with PS1, but was efficiently processed by PS1-dependent γ-secretase. APP-C99 resides in a compartment that is negative for ER, intermediate compartment, and Golgi marker proteins. We conclude that γ-secretase cleavage of APP-C99 occurs in a specialized subcellular compartment where little or no PS1 is detected. This suggests that at least one other factor than PS1, located downstream of the ER, is required for the γ-cleavage of APP-C99. In agreement, we found that intracellular γ-secretase processing of APP-C99-KK both at the γ40 and the γ42 site could be restored partially after brefeldin A treatment. Our data confirm the “spatial paradox” and raise several questions regarding the PS1 is γ-secretase hypothesis.

scholarly journals The odd one out: Arabidopsis reticulon20 has a role in lipid biosynthesis

2017 ◽  
Author(s):  
Verena Kriechbaumer ◽  
Lilly Maneta-Peyret ◽  
Stanley W Botchway ◽  
Jessica Upson ◽  
Louise Hughes ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Sterol Composition ◽  
Lipid Biosynthesis ◽  
Transmembrane Domains ◽  
Eukaryotic Cells ◽  
The Family ◽  
Er Exit Sites ◽  
Punctate Pattern ◽  
Er Membrane

AbstractThe family of reticulon proteins has been shown to be involved in a variety of functions in eukaryotic cells including tubulation of the endoplasmic reticulum (ER), formation of cell plates and primary plasmodesmata. Reticulons are integral ER membrane proteins characterised by a reticulon homology domain comprising four transmembrane domains which results in the reticulons sitting in the membrane in a W-topology. Here we report on a subgroup of reticulons with an extended N-terminal domain and in particular on arabidopsis reticulon 20. We show that reticulon 20 is located in a unique punctate pattern on the ER membrane. Its closest homologue reticulon 19 labels the whole ER. We show that mutants in RTN20 or RTN19, respectively, display a significant change in sterol composition in the roots indicating a role in lipid biosynthesis or regulation. A third homologue in this family - 3BETAHSD/D1- is localised to ER exit sites resulting in an intriguing location difference for the three proteins.

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