The Golgin Coiled-Coil Proteins of the Golgi Apparatus

A number of long coiled-coil proteins are present on the Golgi. Often referred to as “golgins,” they are well conserved in evolution and at least five are likely to have been present in the last common ancestor of all eukaryotes. Individual golgins are found in different parts of the Golgi stack, and they are typically anchored to the membrane at their carboxyl termini by a transmembrane domain or by binding a small GTPase. They appear to have roles in membrane traffic and Golgi structure, but their precise function is in most cases unclear. Many have binding sites for Rab family GTPases along their length, and this has led to the suggestion that the golgins act collectively to form a tentacular matrix that surrounds the Golgi to capture Rab-coated membranes in the vicinity of the stack. Such a collective role might explain the lack of cell lethality seen following loss of some of the genes in human familial conditions or mouse models.Coiled-coils are widely occurring protein structural motifs in which two or more α-helices wind around each other to form an extended rod-like structure. Proteins containing such structures are found in many parts of the cell, and play diverse roles including organizing centrosomes, chromatin, and synapses, or serving as molecular motors. As such there may seem little reason to consider them collectively beyond an interest in the structural and biophysical properties of the coiled-coil itself. However, the Golgi is unique amongst the cellular compartments in that several different large coiled-coil proteins are present on its cytoplasmic surface (Gillingham and Munro 2003; Lupashin and Sztul 2005; Short et al. 2005; Ramirez and Lowe 2009). A number of these share a similar organization in that most of the protein is predicted to form a coiled-coil, and that their carboxyl termini mediate attachment to Golgi membranes. They are generally ubiquitously expressed and well conserved in evolution, but their coiled-coil regions are relatively poorly conserved suggesting that much of their length serves as spacer. Given that 500 residues of coiled-coil is ∼75 nm in length then the proteins could extend for ∼100–400 nm. Some of the proteins have regions which appear likely to be unstructured and hence could serve as extensions or hinges to increase the proteins’ reach and flexibility (Oas and Endow 1994; Yamakawa et al. 1996). These shared features suggest that the proteins serve related functions on the Golgi. The term “golgin” is often applied to these proteins having been coined in early studies when several were found as human autoantigens (Fritzler et al. 1993), but the term lacks a clear definition. To provide a focus to this article, I will concentrate on “golgins” as defined by being a protein that is found primarily, if not exclusively, on the Golgi and is predicted to form a homodimeric parallel coiled-coil over most of its length. Proteins with shorter regions of coiled-coil are more likely to have roles distinct to the golgins, especially if further domains are present.Golgin coiled-coil proteins are found on the cis-face of the Golgi, around the rims of the stack and on the trans-face of the Golgi (Fig. 1). The human golgins are summarized in Open in a separate windowFigure 1.The golgin coiled-coil proteins of humans.Schematic representations of known human golgins. Regions predicted to form coiled-coils are shown in gray, and known domains involved in protein function or subcellular targeting are indicated.

Table 1.

The canonical golgins of the human Golgi and their orthologs.

Golgin160 recruits the dynein motor to position the Golgi apparatus

Membrane motility is a fundamental characteristic of all eukaryotic cells. One of the best-known examples is that of the mammalian Golgi apparatus, where constant inward movement of Golgi membranes results in its characteristic position near the centrosome. While it is clear that the minus-end-directed motor dynein is required for this process, the mechanism and regulation of dynein recruitment to Golgi membranes remains unknown. Here, we show that the Golgi protein golgin160 recruits dynein to Golgi membranes. This recruitment confers centripetal motility to membranes and is regulated by the GTPase Arf1. Further, during cell division, motor association with membranes is regulated by the dissociation of the receptor-motor complex from membranes. These results identify a cell-cycle-regulated membrane receptor for a molecular motor and?suggest a mechanistic basis for achieving the dramatic changes in organelle positioning seen during cell division.     

The intraflagellar transport protein IFT20 is associated with the Golgi complex and is required for cilia assembly

Eukaryotic cilia are assembled via intraflagellar transport (IFT) in which large protein particles are motored along ciliary microtubules. The IFT particles are composed of at least 17 polypeptides that are thought to contain binding sites for various cargos that need to be transported from their site of synthesis in the cell body to the site of assembly in the cilium. We show here that the IFT20 subunit of the particle is localized to the Golgi complex in addition to the basal body and cilia where all previous IFT particle proteins had been found. In living cells, fluorescently tagged IFT20 is highly dynamic and moves between the Golgi complex and the cilium as well as along ciliary microtubules. Strong knock down of IFT20 in mammalian cells blocks ciliary assembly but does not affect Golgi structure. Moderate knockdown does not block cilia assembly but reduces the amount of polycystin-2 that is localized to the cilia. This work suggests that IFT20 functions in the delivery of ciliary membrane proteins from the Golgi complex to the cilium.     

The Golgin Tether Giantin Regulates the Secretory Pathway by Controlling Stack Organization within Golgi Apparatus

Golgins are coiled-coil proteins that play a key role in the regulation of Golgi architecture and function. Giantin, the largest golgin in mammals, forms a complex with p115, rab1, GM130, and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), thereby facilitating vesicle tethering and fusion processes around the Golgi apparatus. Treatment with the microtubule destabilizing drug nocodazole transforms the Golgi ribbon into individual Golgi stacks. Here we show that siRNA-mediated depletion of giantin resulted in more dispersed Golgi stacks after nocodazole treatment than by control treatment, without changing the average cisternal length. Furthermore, depletion of giantin caused an increase in cargo transport that was associated with altered cell surface protein glycosylation. Drosophila S2 cells are known to have dispersed Golgi stacks and no giantin homolog. The exogenous expression of mammalian giantin cDNA in S2 cells resulted in clustered Golgi stacks, similar to the Golgi ribbon in mammalian cells. These results suggest that the spatial organization of the Golgi ribbon is mediated by giantin, which also plays a role in cargo transport and sugar modifications.     

Identification of a novel protein (G210) specific to the Golgi apparatus

A monoclonal antibody, 3C9, has enabled the detection of a novel Golgi-specific protein in bovine tissues. Immunohistochemical studies at the light microscopic level have detected the 3C9 antigen only in certain cells: exocrine pancreas, gut epithelium, and thymus epithelium. Examination of gut and pancreas by immunoelectron microscopy showed a localization exclusive to the Golgi apparatus. The relative molecular weight of the antigen detected by immunoblotting is 210,000 daltons. The antigen is not extracted from microsomal membranes of bovine gut epithelium by sodium carbonate solutions. Furthermore, the 3C9 antigen enters into the detergent phase when Triton X-114 partitioning methods are used. These data strongly suggest that this novel antigen is an intrinsic membrane protein, resident in the Golgi apparatus of certain cells. Moreover, they enhance the hypothesis that the distribution of enzymes and polypeptides in the Golgi apparatus is cell specific.     

The Mre11 Complex and the Response to Dysfunctional Telomeres

In this study, we examine the telomeric functions of the mammalian Mre11 complex by using hypomorphic Mre11 and Nbs1 mutants (Mre11^ATLD1/ATLD1 and Nbs1^ΔB/ΔB, respectively). No telomere shortening was observed in Mre11^ATLD1/ATLD1 cells after extensive passage through culture, and the rate of telomere shortening in telomerase-deficient (Tert^Δ/Δ) Mre11^ATLD1/ATLD1 cells was the same as that in Tert^Δ/Δ alone. Although telomeres from late-passage Mre11^ATLD1/ATLD1 Tert^Δ/Δ cells were as short as those from Tert^Δ/Δ, the incidence of telomere fusions was reduced. This effect on fusions was also evident upon acute telomere dysfunction in Mre11^ATLD1/ATLD1 and Nbs1^ΔB/ΔB cells rendered Trf2 deficient by cre-mediated TRF2 inactivation than in wild-type cells. The residual fusions formed in Mre11 complex mutant cells exhibited a strong tendency toward chromatid fusions, with an almost complete bias for fusion of telomeres replicated by the leading strand. Finally, the response to acute telomere dysfunction was strongly impaired by Mre11 complex hypomorphism, as the formation of telomere dysfunction-induced DNA damage foci was reduced in both cre-infected Mre11^ATLD1/ATLD1 Trf2^F/Δ and Nbs1^ΔB/ΔB Trf2^F/F cells. These data indicate that the Mre11 complex influences the cellular response to telomere dysfunction, reminiscent of its influence on the response to interstitial DNA breaks, and suggest that it may promote telomeric DNA end processing during DNA replication.The Mre11 complex (in mammals, Mre11, Rad50, and Nbs1) plays a central role in the cellular response to DNA double-strand breaks (DSBs). The Mre11 complex acts as a DSB sensor, promoting the activation of ATM-dependent DNA damage signaling pathways, DNA repair, and apoptosis. In addition, the complex plays a direct role in recombinational DNA repair, influencing both homologous recombination and nonhomologous end joining (NHEJ) (39). The Mre11 complex''s diverse functions in the DNA damage response are likely predicated on its physical association with chromatin. In this regard, one of the least-understood roles of the Mre11 complex in mammals is its association with telomeres.In mammals, telomeric DNA consists of double-stranded TTAGGG repeats ending in a single-stranded 3′ G overhang, and an array of telomere binding proteins called the shelterin complex that function to prevent telomeres from being recognized as DNA breaks (33). DNA of the overhang invades the double-stranded telomeric repeat sequence to form a t-loop structure (14, 32). The formation of the t-loop requires the telomere protection and remodeling proteins that make up the shelterin complex (7), and these may also contribute to telomere length regulation by preventing telomerase access to chromosomal ends.Data regarding the role of the Mre11 complex at the telomere have implicated the Mre11 complex in several aspects of telomere maintenance and function. For example, it has been suggested that the Mre11 complex may promote formation of the 3′ telomeric overhang by influencing 5′-to-3′ resection of newly replicated chromosome ends (6). In Saccharomyces cerevisiae, the Mre11 complex recruits the ATM orthologue, Tel1, which is in turn required to recruit telomerase (12, 45). Consequently, Mre11 complex deficiency results in telomere shortening. In mammals, recruitment of telomerase is thought to be regulated primarily by the telomeric protein components TRF1, TPP1, and POT1 (24, 46, 53). However, telomere shortening has also been noted to occur in cell lines from Nijmegen breakage syndrome (NBS) patients in which a hypomorphic Nbs1 allele is expressed, leading to the suggestion that the Mre11 complex may also promote telomerase function in mammals (36). The Mre11 complex associates with telomeres through its interaction with the shelterin component Trf2, apparently in a cell cycle-dependent manner (47, 54). The significance of this physical association is unclear, as genetic depletion of Rad50, a component of the Mre11 complex, does not phenocopy depletion of Trf2 in most respects (1).To examine the function of the Mre11 complex at mammalian telomeres, we established mouse embryonic fibroblasts (MEFs) derived from a mouse expressing the hypomorphic Mre11^ATLD1 allele, crossed to telomerase deficient Tert^Δ/Δ mice (23, 42), and assessed the rate of telomere shortening. Mre11 complex hypomorphism in MEFs did not affect telomere length, irrespective of telomerase status. In Mre11^ATLD1/ATLD1 Tert^Δ/Δ cells, the fusion of eroded telomeres was reduced compared to Tert^Δ/Δ cells with telomeres shortened to the same extent, suggesting that the Mre11 complex is involved in the response to critically short telomeres. This interpretation was supported by data obtained using a conditional Trf2 allele to generate acute telomere dysfunction in Mre11^ATLD1/ATLD1 and Nbs1^ΔB/ΔB cells. Collectively the data support a role for the Mre11 complex in the recognition and signaling of dysfunctional telomeres. The character of fusions arising in cre-infected Mre11^ATLD1/ATLD1 Trf2^F/Δ and Nbs1^ΔB/ΔB Trf2^F/F cells further suggests that the Mre11 complex may influence the processing of chromosome ends following DNA replication en route to t-loop formation.     

The overexpression of GMAP-210 blocks anterograde and retrograde transport between the ER and the Golgi apparatus

Golgi Microtubule-Associated Protein (GMAP)-210 is a peripheral coiled-coil protein associated with the cis -Golgi network that interacts with microtubule minus ends. GMAP-210 overexpression has previously been shown to perturb the microtubule network and to induce a dramatic enlargement and fragmentation of the Golgi apparatus (Infante C, Ramos-Morales F, Fedriani C, Bornens M, Rios RM. J Cell Biol 1999; 145: 83–98). We now report that overexpressing GMAP-210 blocks the anterograde transport of both a soluble form of alkaline phosphatase and the hemagglutinin protein of influenza virus, an integral membrane protein, between the endoplasmic reticulum and the cis /medial (mannosidase II-positive) Golgi compartment. Retrograde transport of the Shiga toxin B-subunit is also blocked between the Golgi apparatus and the endoplasmic reticulum. As a consequence, the B-subunit accumulates in compartments positive for GMAP-210. Ultrastructural analysis revealed that, under these conditions, the Golgi complex is totally disassembled and Golgi proteins as well as proteins of the intermediate compartment are found in vesicle clusters distributed throughout the cell. The role of GMAP-210 on membrane processes at the interface between the endoplasmic reticulum and the Golgi apparatus is discussed in the light of the property of this protein to bind CGN membranes and microtubules.     

Mutant Huntingtin Impairs Post-Golgi Trafficking to Lysosomes by Delocalizing Optineurin/Rab8 Complex from the Golgi Apparatus

Huntingtin regulates post-Golgi trafficking of secreted proteins. Here, we studied the mechanism by which mutant huntingtin impairs this process. Colocalization studies and Western blot analysis of isolated Golgi membranes showed a reduction of huntingtin in the Golgi apparatus of cells expressing mutant huntingtin. These findings correlated with a decrease in the levels of optineurin and Rab8 in the Golgi apparatus that can be reverted by overexpression of full-length wild-type huntingtin. In addition, immunoprecipitation studies showed reduced interaction between mutant huntingtin and optineurin/Rab8. Cells expressing mutant huntingtin produced both an accumulation of clathrin adaptor complex 1 at the Golgi and an increase of clathrin-coated vesicles in the vicinity of Golgi cisternae as revealed by electron microscopy. Furthermore, inverse fluorescence recovery after photobleaching analysis for lysosomal-associated membrane protein-1 and mannose-6-phosphate receptor showed that the optineurin/Rab8-dependent post-Golgi trafficking to lysosomes was impaired in cells expressing mutant huntingtin or reducing huntingtin levels by small interfering RNA. Accordingly, these cells showed a lower content of cathepsin D in lysosomes, which led to an overall reduction of lysosomal activity. Together, our results indicate that mutant huntingtin perturbs post-Golgi trafficking to lysosomal compartments by delocalizing the optineurin/Rab8 complex, which, in turn, affects the lysosomal function.     

ENTH/ANTH domains expand to the Golgi

Clathrin-coated vesicles (CCVs) play important roles in nutrient uptake, downregulation of signaling receptors, pathogen invasion and biogenesis of endosomes and lysosomes. Although detailed models for endocytic CCV formation have emerged, the process of CCV formation at the Golgi and endosomes has been less clear. Key to endocytic CCV formation are proteins containing related phosphoinositide-binding ENTH and ANTH domains. Now, recent studies have identified novel ENTH/ANTH proteins that participate in CCV-mediated traffic between the trans-Golgi Network (TGN) and endosomes and have defined a molecular basis for interaction with AP-1 and GGA adaptors in clathrin coats of the TGN/endosomes. Thus, ENTH/ANTH domain proteins appear to be universal elements in nucleation of clathrin coats.     

The Highly Conserved COPII Coat Complex Sorts Cargo from the Endoplasmic Reticulum and Targets It to the Golgi

Protein egress from the endoplasmic reticulum (ER) is driven by a conserved cytoplasmic coat complex called the COPII coat. The COPII coat complex contains an inner shell (Sec23/Sec24) that sorts cargo into ER-derived vesicles and an outer cage (Sec13/Sec31) that leads to coat polymerization. Once released from the ER, vesicles must tether to and fuse with the target membrane to deliver their protein and lipid contents. This delivery step also depends on the COPII coat, with coat proteins binding directly to tethering and regulatory factors. Recent findings have yielded new insight into how COPII-mediated vesicle traffic is regulated. Here we discuss the molecular basis of COPII-mediated ER–Golgi traffic, focusing on the surprising complexity of how ER-derived vesicles form, package diverse cargoes, and correctly target these cargoes to their destination.The port of entry into the secretory pathway is the endoplasmic reticulum (ER). Approximately one-third of the eukaryotic proteome traffics from this multifunctional organelle (Huh et al. 2003). This diverse set of cargo is translocated into the ER, folded, and modified before it travels to the Golgi, where further modifications occur. From the Golgi, cargo is sorted to other subcellular compartments to perform a variety of cellular functions. The highly conserved machinery required for these transport events was initially identified through genetic screens in the yeast Saccharomyces cerevisiae, and insights into the function of this machinery were provided through the use of in vitro transport assays. Advances in microscopy, in particular, the use of GFP fusion proteins and live cell imaging, have also played a critical role in understanding the dynamics of membrane traffic. In this article, we describe the mechanistic advances that have helped us to understand how diverse cargo correctly traffics from the ER to the Golgi complex in lower and higher eukaryotes. Even though these mechanisms are largely conserved, they are more complex at the molecular and organizational levels in metazoans.     

GM1 Synthase Depends on N-Glycosylation for Enzyme Activity and Trafficking to the Golgi Complex

Glycosyltransferase cDNAs contain a variable number of potential N-glycosylation sites. Here we examined the occupancy and relevance for the activity and intracellular trafficking of the only potential N-glycosylation site of the mouse 1,3galactosyltransferase (Gal-T2 or GA1/GM1/GD1b synthase) in Gal-T2 cDNA transfected CHO-K1 cells. Transfected cells synthesize a Golgi located active enzyme of 43 kDa whose N-glycan was metabolically labeled from [³H]mannose and was Endo-H sensitive. Inhibition of N-glycosylation by Tunicamycin or by point mutation of the N-glycosylation site resulted in the synthesis of a polypeptide of 40 kDa which lacked enzyme activity and was concentrated in the endoplasmic reticulum (ER). Inhibition of ER glucosidases by Castanospermine impaired the exit of a form of Gal-T2 having reduced enzyme activity from the ER. The N-terminal Gal-T2 domain (aa 1–52) was able to direct and to retain the green fluorescence protein in the Golgi complex. Taken together, these results indicate that Gal-T2 depends on N-glycosylation for its activity and for proper trafficking to, but not its retention in, the Golgi complex.     

An Integrated Physical Map of 210 Markers Assigned to the Short Arm of Human Chromosome 11

Using a panel of patient cell lines with chromosomal breakpoints, we constructed a physical map for the short arm of human chromosome 11. We focused on 11p15, a chromosome band harboring at least 25 known genes and associated with the Beckwith-Wiedemann syndrome, several childhood tumors, and genomic imprinting. This underlines the need for a physical map for this region. We divided the short arm of chromosome 11 into 18 breakpoint regions, and a large series of new and previously described genes and markers was mapped within these intervals using fluorescence in situ hybridization. Cosmid fingerprint analysis showed that 19 of these markers were included in cosmid contigs. A detailed 10-Mb pulsed-field physical map of the region 11p15.3-pter was constructed. These three different approaches enabled the high-resolution mapping of 210 markers, including 22 known genes.     

Conserved Motif of CDK5RAP2 Mediates Its Localization to Centrosomes and the Golgi Complex

As the primary microtubule-organizing centers, centrosomes require γ-tubulin for microtubule nucleation and organization. Located in close vicinity to centrosomes, the Golgi complex is another microtubule-organizing organelle in interphase cells. CDK5RAP2 is a γ-tubulin complex-binding protein and functions in γ-tubulin attachment to centrosomes. In this study, we find that CDK5RAP2 localizes to the Golgi complex in an ATP- and centrosome-dependent manner and associates with Golgi membranes independently of microtubules. CDK5RAP2 contains a centrosome-targeting domain with its core region highly homologous to the Motif 2 (CM2) of centrosomin, a functionally related protein in Drosophila. This sequence, referred to as the CM2-like motif, is also conserved in related proteins in chicken and zebrafish. Therefore, CDK5RAP2 may undertake a conserved mechanism for centrosomal localization. Using a mutational approach, we demonstrate that the CM2-like motif plays a crucial role in the centrosomal and Golgi localization of CDK5RAP2. Furthermore, the CM2-like motif is essential for the association of the centrosome-targeting domain to pericentrin and AKAP450. The binding with pericentrin is required for the centrosomal and Golgi localization of CDK5RAP2, whereas the binding with AKAP450 is required for the Golgi localization. Although the CM2-like motif possesses the activity of Ca²⁺-independent calmodulin binding, binding of calmodulin to this sequence is dispensable for centrosomal and Golgi association. Altogether, CDK5RAP2 may represent a novel mechanism for centrosomal and Golgi localization.     

The Endosomal Pathway and the Golgi Complex Are Involved in the Infectious Bursal Disease Virus Life Cycle

Infectious bursal disease virus (IBDV), a double-stranded RNA virus belonging to the Birnaviridae family, causes immunosuppression in chickens. In this study, we defined the localization of IBDV replication complexes based on colocalization analysis of VP3, the major protein component of IBDV ribonucleoproteins (RNPs). Our results indicate that VP3 localizes to vesicular structures bearing features of early and late endocytic compartments located in the juxtanuclear region. Interfering with the endocytic pathway with a dominant negative version of Rab5 after the internalization step leads to a reduction in virus titer. Triple-immunostaining studies between VP3, the viral RNA-dependent RNA polymerase VP1, and viral double-stranded RNA (dsRNA) showed a well-defined colocalization, indicating that the three critical components of the RNPs colocalize in the same structure, likely representing replication complexes. Interestingly, recombinant expressed VP3 also localizes to endosomes. Employing Golgi markers, we found that VP3-containing vesicles were closely associated with this organelle. Depolymerization of microtubules with nocodazole caused a profound change in VP3 localization, showing a punctate distribution scattered throughout the cytoplasm. However, these VP3-positive structures remained associated with Golgi ministacks. Similarly, brefeldin A (BFA) treatment led to a punctate distribution of VP3, scattered throughout the cytoplasm of infected cells. In addition, analysis of intra- and extracellular viral infective particles after BFA treatment of avian cells suggested a role for the Golgi complex in viral assembly. These results constitute the first study elucidating the localization of IBDV replication complexes (i.e., in endocytic compartments) and establishing a role for the Golgi apparatus in the assembly step of a birnavirus.     

Proteolytic processing of pro-ACTH/endorphin begins in the Golgi complex of pituitary corticotropes and AtT-20 cells

The intracellular sites where proteolytic processing of pro-ACTH/endorphin or POMC take place have not yet been reliably identified. We have used affinity-purified antisera that recognize only the products of POMC processing and immunoelectron microscopy to identify the compartments of rat pituitary corticotropes and mouse AtT-20 cells in which cleavage occurs. Immunoperoxidase labeling of cryostat sections and immunogold labeling of ultrathin frozen sections were used for localization of the processing sites. By both procedures we detected processed peptides in Golgi cisternae and secretion granules. Within the Golgi, labeling was limited to the last or transmost cisterna and was most concentrated in its dilated rims which contain condensing secretory protein. No labeling of other Golgi cisternae was seen. All Golgi cisternae were labeled, however, when antisera that recognize unprocessed POMC were used for immunolabeling. We conclude that in AtT-20 and rat pituitary cells: 1) processing of POMC through at least two endo- and exoproteolytic cleavage steps and alpha-amidation of joining peptide begin in the trans Golgi subcompartment; 2) no detectable processing takes place before POMC reaches the trans Golgi cisterna; and 3) this Golgi cisterna as well as secretion granules contain the active enzymes necessary for proteolytic processing and alpha-amidation.     

Nonclathrin Vesicle Coats and Filament Networks in the Transition Zone and trans-Golgi Region of the Golgi Complex of Paramecium

Understanding vesicle trafficking to and through the Golgi stack has been greatly elucidated recently, but the question of what holds the endoplasmic reticulum (ER) and Golgi stack together in many cell types and an explanation of anterograde trafficking in the ER-Golgi transitional zone have not yet been adequately explained. We have studied these problems using both the thin sectioning and the quick-freeze deep-etch (QF-DE) technique on Paramecium cells harvested at different culture ages. Although the Golgi apparatus of Paramecium is made up of many sets of more reduced stacks of cisternae than those of many mammalian cells, the stacks in Paramecium always bear a close relationship to a transitional element of the ER from which non-clathrin-coated transition vesicles arise. In QF-DE replicas two networks of filaments are clearly shown; one is in this ER-Golgi transition zone and the other is on the trans side of the Golgi stack. The network associated with the trans-Golgi region links a number of vesicular elements. The network in the transition zone spans the distance between the ER and the cis-cisterna of the Golgi stack and has branches extending to the coats of the enmeshed nonclathrin-coated transition vesicles. These coats consist of a layer of 11-nm globular elements (the same size as coatomer complexes) which surround the 40-nm-diameter transition vesicles. We conclude that the filamentous network holds the ER and Golgi stack together and prevents the dispersal of the transition vesicles away from this zone. This network may also delineate and stabilize the transitional element within the ER and, finally, help organize anterograde transition vesicle trafficking in this ER-Golgi transition zone.