Guanine nucleotides modulate the effects of brefeldin A in semipermeable cells: regulation of the association of a 110-kD peripheral membrane protein with the Golgi apparatus

The release of a 110-kD peripheral membrane protein from the Golgi apparatus is an early event in brefeldin A (BFA) action, preceding the movement of Golgi membrane into the ER. ATP depletion also causes the reversible redistribution of the 110-kD protein from Golgi membrane into the cytosol, although no Golgi disassembly occurs. To further define the effects of BFA on the association of the 110-kD protein with the Golgi apparatus we have used filter perforation techniques to produce semipermeable cells. All previously observed effects of BFA, including the rapid redistribution of the 110-kD protein and the movement of Golgi membrane into the ER, could be reproduced in the semipermeable cells. The role of guanine nucleotides in this process was investigated using the nonhydrolyzable analogue of GTP, GTP gamma S. Pretreatment of semipermeable cells with GTP gamma S prevented the BFA-induced redistribution of the 110-kD protein from the Golgi apparatus and movement of Golgi membrane into the ER. GTP gamma S could also abrogate the observed release of the 110-kD protein from Golgi membranes which occurred in response to ATP depletion. Additionally, when the 110-kD protein had first been dissociated from Golgi membranes by ATP depletion, GTP gamma S could restore Golgi membrane association of the 110-kD protein, but not if BFA was present. All of these effects observed with GTP gamma S in semipermeable cells could be reproduced in intact cells treated with AlF4-. These results suggest that guanine nucleotides regulate the dynamic association/dissociation of the 110-kD protein with the Golgi apparatus and that BFA perturbs this process by interfering with the association of the 110-kD protein with the Golgi apparatus.     

Dissociation of a 110-kD peripheral membrane protein from the Golgi apparatus is an early event in brefeldin A action

Brefeldin A (BFA) has a profound effect on the structure of the Golgi apparatus, causing Golgi proteins to redistribute into the ER minutes after drug treatment. Here we describe the dissociation of a 110-kD cytoplasmically oriented peripheral membrane protein (Allan, V. J., and T. E. Kreis. 1986. J. Cell Biol. 103:2229-2239) from the Golgi apparatus as an early event in BFA action, preceding other morphologic changes. In contrast, other peripheral membrane proteins of the Golgi apparatus were not released but followed Golgi membrane into the ER during BFA treatment. The 110-kD protein remained widely dispersed throughout the cytoplasm during drug treatment, but upon removal of BFA it reassociated with membranes during reformation of the Golgi apparatus. Although a 30-s exposure to the drug was sufficient to cause the redistribution of the 110-kD protein, removal of the drug after this short exposure resulted in the reassociation of the 110-kD protein and no change in Golgi structure. If cells were exposed to BFA for 1 min or more, however, a portion of the Golgi membrane was committed to move into and out of the ER after removal of the drug. ATP depletion also caused the reversible release of the 110-kD protein, but without Golgi membrane redistribution into the ER. These findings suggest that the interaction between the 110-kD protein and the Golgi apparatus is dynamic and can be perturbed by metabolic changes or the drug BFA.     

A brefeldin A-like phenotype is induced by the overexpression of a human ERD-2-like protein, ELP-1.

Brefeldin A (BFA) is a unique drug affecting the molecular mechanisms that regulate membrane traffic and organelle structure. BFA's ability to alter retrograde traffic from the Golgi to the endoplasmic reticulum (ER) led us to ask whether the ERD-2 retrieval receptor, proposed to return escaped ER resident proteins from the Golgi, might either interfere with or mimic the effects of the drug. When either human ERD-2 or a novel human homolog (referred to as ELP-1) is overexpressed in a variety of cell types, the effects are phenotypically indistinguishable from the addition of BFA. These include the redistribution of the Golgi coat protein, beta-COP, to the cytosol, the loss of the Golgi apparatus as a distinct organelle, the mixing of this organelle with the ER, the addition of complex oligosaccharides to resident ER glycoproteins, and the block of anterograde traffic. Thus, these receptors may provide signals that regulate retrograde traffic between the Golgi and the ER.     

Brefeldin A's effects on endosomes, lysosomes, and the TGN suggest a general mechanism for regulating organelle structure and membrane traffic.

Addition of brefeldin A (BFA) to most cells results in both the formation of extensive, uncoated membrane tubules through which Golgi components redistribute into the ER and the failure to transport molecules out of this mixed ER/Golgi system. In this study we provide evidence that suggests BFA's effects are not limited to the Golgi apparatus but are reiterated throughout the central vacuolar system. Addition of BFA to cells resulted in the tubulation of the endosomal system, the trans-Golgi network (TGN), and lysosomes. Tubule formation of these organelles was specific to BFA, shared near identical pharmacologic characteristics as Golgi tubules and resulted in targeted membrane fusion. Analogous to the mixing of the Golgi with the ER during BFA treatment, the TGN mixed with the recycling endosomal system. This mixed system remained functional with normal cycling between plasma membrane and endosomes, but traffic between endosomes and lysosomes was impaired.     

100-kD proteins of Golgi- and trans-Golgi network-associated coated vesicles have related but distinct membrane binding properties

The 100-110-kD proteins (alpha-, beta-, beta'-, and gamma-adaptins) of clathrin-coated vesicles and the 110-kD protein (beta-COP) of the nonclathrin-coated vesicles that mediate constitutive transport through the Golgi have homologous protein sequences. To determine whether homologous processes are involved in assembly of the two types of coated vesicles, the membrane binding properties of their coat proteins were compared. After treatment of MDBK cells with the fungal metabolite Brefeldin A (BFA), beta-COP was redistributed to the cytoplasm within 15 s, gamma-adaptin and clathrin in the trans-Golgi network (TGN) dispersed within 30 s, but the alpha-adaptin and clathrin present on coated pits and vesicles derived from the plasma membrane remained membrane associated even after a 15-min exposure to BFA. In PtK1 cells and MDCK cells, BFA did not affect beta-COP binding or Golgi morphology but still induced redistribution of gamma-adaptin and clathrin from TGN membranes to the cytoplasm. Thus BFA affects the binding of coat proteins to membranes in the Golgi region (Golgi apparatus and TGN) but not plasma membranes. However, the Golgi binding interactions of beta-COP and gamma-adaptin are distinct and differentially sensitive to BFA. BFA treatment did not release gamma-adaptin or clathrin from purified clathrin-coated vesicles, suggesting that their distribution to the cytoplasm after BFA treatment of cells was due to interference with their rebinding to TGN membranes after a normal cycle of disassembly. This was confirmed using an in vitro assay in which gamma-adaptin binding to TGN membranes was blocked by BFA and enhanced by GTP gamma S, similar to the binding of beta-COP to Golgi membranes. These results suggest the involvement of GTP-dependent proteins in the association of the 100-kD coat proteins with membranes in the Golgi region of the cell.     

Gap junction turnover, intracellular trafficking, and phosphorylation of connexin43 in brefeldin A-treated rat mammary tumor cells

Intercellular gap junction channels are thought to form when oligomers of connexins from one cell (connexons) register and pair with connexons from a neighboring cell en route to forming tightly packed arrays (plaques). In the current study we used the rat mammary BICR-M1Rk tumor cell line to examine the trafficking, maturation, and kinetics of connexin43 (Cx43). Cx43 was conclusively shown to reside in the Golgi apparatus in addition to sites of cell-cell apposition in these cells and in normal rat kidney cells. Brefeldin A (BFA) blocked Cx43 trafficking to the surface of the mammary cells and also prevented phosphorylation of the 42-kD form of Cx43 to 44- and 46-kD species. However, phosphorylation of Cx43 occurred in the presence of BFA while it was still a resident of the ER or Golgi apparatus yielding a 43-kD form of Cx43. Moreover, the 42- and 43-kD forms of Cx43 trapped in the ER/Golgi compartment were available for gap junction assembly upon the removal of BFA. Mammary cells treated with BFA for 6 h lost preexisting gap junction "plaques," as well as the 44- and 46-kD forms of Cx43 and functional coupling. These events were reversible 1 h after the removal of BFA and not dependent on protein synthesis. In summary, we provide strong evidence that in BICR-M1Rk tumor cells: (a) Cx43 is transiently phosphorylated in the ER/Golgi apparatus, (b) Cx43 trapped in the ER/Golgi compartment is not subject to rapid degradation and is available for the assembly of new gap junction channels upon the removal of BFA, (c) the rapid turnover of gap junction plaques is correlated with the loss of the 44- and 46-kD forms of Cx43.     

Involvement of the Golgi region in the intracellular trafficking of cholera toxin

The intracellular pathway following receptor-mediated endocytosis of cholera toxin was studied using brefeldin A (BFA), which inhibited protein secretion and induced dramatic morphological changes in the Golgi region. In both mouse Y1 adrenal cells and CHO cells, BFA at 1 μg/ml caused a 80–90% inhibition of the cholera toxin (CT)-elevation of intracellular cAMP. The inhibition of the cytotoxicity of CT by BFA was also observed in a rounding assay of Y1 adrenal cells. The inhibition of CT cytotoxicity by BFA was dose dependent, with the ID₅₀ value similar to the LD₅₀ of BFA in Y1 adrenal cells. Binding and internalization of [¹²⁵I]-cholera toxin in Y1 adrenal cells was not affected by BFA. Unlike the BFA-sensitive cell lines such as Y1 adrenal and CHO cells, BFA at 1 μg/ml did not inhibit the cytotoxicity of CT in PtK₁ cells, of which the Golgi structure was BFA-resistant. These results strongly suggest that a BFA-sensitive Golgi is required for the protection of CT cytotoxicity by BFA. In contrast, elevation of the intracellular cAMP by forskolin, which acts directly on the plasma membrane adenylate cyclase, was not affected by BFA. These observations indicate that the intoxication of target cells by CT requires an intact Golgi region for its intracellular trafficking and/or processing. In this respect, CT shares a common intracellular pathway with ricin, Pseudomonas toxin, and modeccin, even though their structures and modes of action are very different. © 1993 Wiley-Liss, Inc.     

Structural integrity of the Golgi stack is essential for normal secretory functions of rat parotid acinar cells: effects of brefeldin A and okadaic acid.

We examined the effects of specific inhibitors, brefeldin A (BFA) and okadaic acid (OA), on the ultrastructural organization of the Golgi apparatus and distributions of amylase, Golgi-associated proteins, and cathepsin D in the rat parotid acinar cells. BFA induced a rapid regression of the Golgi stack into rudimentary Golgi clusters composed of tubulovesicules, in parallel with a redistribution of the Golgi-resident proteins and a coat protein (beta-COP) into the region of the rough endoplasmic reticulum (rER) or cytosol. The rapid disruption of the Golgi stack could also be induced by the effect of OA. However, redistribution of the Golgi proteins in rER or cytosol could not be observed and beta-COP was not dispersed but was retained on the rudimentary Golgi apparatus. These findings suggested that the mechanism of OA in inducing degeneration of the Golgi stack was markedly different from that of BFA. In addition, missorting of amylase, a Golgi protein, and cathepsin D into incorrect transport pathways is apparent in the course of the disruption of the Golgi stack by OA. These Golgi-disrupting effects are reversible and the reconstruction of the stacked structure of the Golgi apparatus started immediately after the removal of inhibitors. In the recovery processes, missorting was also observed until the integrated structure of the Golgi apparatus was completely reconstructed. This suggested that the integrated structure of the Golgi apparatus was quite necessary for the occurrence of normal secretory events, including proper sorting of molecules.     

Isolation and characterization of a brefeldin A-resistant mutant of monkey kidney Vero cells.

Brefeldin A (BFA) is a fungal antibiotic which disrupts protein transport between the endoplasmic reticulum and the Golgi. A BFA-resistant mutant of monkey kidney Vero cells, BER-40, which exhibited about a 90-fold increase in the LD50 of BFA (5.2 ng/ml for Vero cells versus 460 ng/ml for BER-40 cells), has been isolated. The increased resistance of BER-40 cells toward BFA was also manifested in a greatly reduced inhibition of protein secretion by BFA in the mutant and a lack of protection by BFA of the mutant cells from ricin cytotoxicity. Somatic cell hybridization between the Vero and BER-40 cells showed that the BFA-resistance in BER-40 behaved as a codominant trait. The structure of the Golgi region, as examined by immunofluorescence microscopy with antibodies against Golgi markers (the 110-kDa protein and mannosidase II) or with fluorescent lipid NBD-ceramide, was unchanged in the mutant cells as compared to that in the wild-type cells. Treatment of Vero cells with BFA (1 micrograms/ml) or with 2-deoxyglucose plus sodium azide resulted in a rapid release of the 110-kDa protein, mannosidase II, and NBD-ceramide from the Golgi membrane to a more diffuse distribution in the cytosol. In contrast, these three Golgi markers remained to be Golgi-associated following treatment of BER-40 cells with BFA or with 2-deoxyglucose plus sodium azide. Immunoblotting of cell extracts from Vero and BER-40 cells with monoclonal antibody against the 110-kDa protein did not reveal any significant difference in the level of this Golgi marker in the mutant cells. These data suggest that the BFA-resistance mutation in BER-40 has rendered the cyclic pathway of the 110-kDa protein assembly to the Golgi membrane resistant to both BFA and 2-deoxyglucose plus sodium azide.     

Stimulation of Postsynaptic and Inhibition of Presynaptic Adenylyl Cyclase Activity by Metabotropic Glutamate: Receptor Activation

Abstract: To determine the subcellular distribution of cyclic AMP-coupled metabotropic glutamate receptors (mGluRs), the effects of glutamate agonists on adenylyl cyclase activity were examined using two hippocampal membrane preparations. These were synaptosomes (SY), which are composed of presynaptic terminals, and synaptoneurosomes (SN), which are composed of both pre-and postsynaptic elements. In SY, a water-soluble analogue of forskolin (7β-forskolin) increased enzyme activity ˜ 10-fold at the highest concentration tested. The selective metabotropic receptor agonist (1S,3 R )-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3 R -ACPD) inhibited enzyme activity as did glutamate and quisqualate. l -Amino-4-phosphobutanoate ( l -AP4) had no effect on enzyme activity at any concentration tested. The metabotropic receptor antagonist l -2-amino-3-phosphopropionic acid ( l -AP3) was not effective in the SY in antagonizing the agonist-induced decreases in adenylyl cyclase activity by glutamate or 1S,3 R -ACPD. It was, however, effective at antagonizing quisqualate-induced decreases in enzyme activity. In SN, at the highest concentration tested, 7β-forskolin produced a 60-fold increase in adenylyl cyclase activity. As was observed in SY, glutamate decreased adenylyl cyclase activity in SN. In contrast, 1S,3 R -ACPD, quisqualate, and l -AP4 increased adenylyl cyclase activity. In the SN, l -AP3 was ineffective in antagonizing any agonist-induced increases (1S,3 R -ACPD, l -AP4, and quisqualate) or decreases (glutamate) in adenylyl cyclase activity. The data suggest that postsynaptic metabotropic glutamate receptor activation results in stimulation of adenylyl cyclase activity, whereas inhibition of this enzyme appears to be mediated at least partly through presynaptic mechanisms.     

Pulmonary surfactant phosphatidylcholine transport bypasses the brefeldin A sensitive compartment of alveolar type II cells

Brefeldin A (BFA) causes disassembly of the Golgi apparatus and blocks protein transport to this organelle from the endoplasmic reticulum. However, there still remains considerable ambiguity regarding the involvement of the Golgi apparatus in glycerolipid transport pathways. We examined the effects of BFA upon the intracellular translocation of phosphatidylcholine in alveolar type II cells, that synthesize, transport, store and secrete large amounts of phospholipid for regulated exocytosis. BFA at concentrations as high as 10 microg/ml failed to alter the assembly of phosphatidylcholine into lamellar bodies, the specialized storage organelles for pulmonary surfactant. The same concentration of BFA was also ineffective at altering the secretion of newly synthesized phosphatidylcholine from alveolar type II cells. In contrast, concentrations of the drug of 2.5 microg/ml completely arrested newly synthesized lysozyme secretion from the same cells, indicating that BFA readily blocked protein transport processes in alveolar type II cells. The disassembly of the Golgi apparatus in alveolar type II cells following BFA treatment was also demonstrated by showing the redistribution of the resident Golgi protein MG-160 to the endoplasmic reticulum. These results indicate that intracellular transport of phosphatidylcholine along the secretory pathway in alveolar type II cells proceeds via a BFA insensitive route and does not require a functional Golgi apparatus.     

G_(i/o)蛋白介导的信息转导通路在低氧预处理心肌保护中的作用

实验以低氧 3h后复氧期间心肌细胞的生存率和LDH的释放量为指标 ,观察Gi/o蛋白及其下游成分在低氧预处理 (hypoxicpreconditioning ,HP)心肌保护中的作用。与单纯低氧组相比 ,HP组 ( 2 5min低氧 30min复氧作为HP)细胞生存率增高 ,LDH释放减少 (P <0 0 1)。用NEM预处理 ,能完全模拟HP的心肌细胞保护作用 ;而用PTX阻断Gi/o蛋白 ,或Forskolin和 8 Br cAMP预处理后 ,再给予HP及低氧 3h/复氧 1h ,则细胞生存率降低 ,LDH释放增加 (P <0 0 1) ;U 7312 2预处理后 ,细胞生存率和LDH释放量无差异 (P >0 0 5 )。结果提示 :Gi/o蛋白通过抑制AC ,减少第二信使cAMP的生成介导了HP的心肌保护作用。PLC可能不参与HP的心肌保护作用     

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

Brefeldin A (BFA), a fungal fatty acid derivative, is a potentagent for disrupting the Golgi apparatus in plant and animalcells. We have examined its action using marker antibodies whichrecognize an epitope in the plant Golgi apparatus (JIM 84),and for proteins held in the endoplasmic reticulum by the HDELER-retention signal (2E7), in combination with double immunolabelling.In maize root cells, disruption of the ER occurs after breakdownof the Golgi apparatus is initiated. The redistribution of theGolgi is shown to be predominantly separate from that of theER, and as with the Golgi, the action of BFA on the ER is alsoreversible. The mode of action of BFA on the ER and Golgi ofplant cells is compared with that described for animal cells. Key words: Zea mays L, Brefeldin A, plant cells, endoplasmic reticulum, Golgi apparatus     

Identification of a 200-kD, brefeldin-sensitive protein on Golgi membranes

A mAb AD7, raised against canine liver Golgi membranes, recognizes a novel, 200-kD protein (p200) which is found in a wide variety of cultured cell lines. Immunofluorescence staining of cultured cells with the AD7 antibody produced intense staining of p200 in the juxtanuclear Golgi complex and more diffuse staining of p200 in the cytoplasm. The p200 protein in the Golgi complex was colocalized with other Golgi proteins, including mannosidase II and beta-COP, a coatomer protein. Localization of p200 by immunoperoxidase staining at the electron microscopic level revealed concentrations of p200 at the dilated rims of Golgi cisternae. Biochemical studies showed that p200 is a peripheral membrane protein which partitions to the aqueous phase of Triton X-114 solutions and is phosphorylated. The p200 protein is located on the cytoplasmic face of membranes, since it was accessible to trypsin digestion in microsomal preparations, and is recovered in approximately equal amounts in membrane pellets and in the cytosol of homogenized cells. Immunofluorescence staining of normal rat kidney cells exposed to the toxin brefeldin A (BFA), showed that there was very rapid redistribution of p200, which was dissociated from Golgi membranes in the presence of this drug. The effect of BFA was reversible, since upon removal of the toxin, AD7 rapidly reassociated with the Golgi complex. In the BFA-resistant cell line PtK1, BFA failed to cause redistribution of p200 from Golgi membranes. Taken together, these results indicate that the p200 Golgi membrane-associated protein has many properties in common with the coatomer protein, beta-COP.     

Golgi proteins persist in the tubulovesicular remnants found in brefeldin A-treated pancreatic acinar cells.

Brefeldin A (BFA) blocks protein export from the endoplasmic reticulum (ER) and causes dismantling of the Golgi cisternae with relocation of resident Golgi proteins to the ER in many cultured cell lines. We examined the effects of BFA on Golgi organization and the distribution of Golgi markers in the rat exocrine pancreas. Immediately after BFA addition, Golgi stacks began to disorganize and Golgi cisternae to vesiculate, and by 15 min no intact Golgi cisternae remained. However, even after prolonged BFA incubation, clusters of small vesicles surrounded by transitional elements of the ER persisted both in the Golgi region and dispersed throughout the apical cytoplasm. These vesicles were morphologically heterogeneous in the density of their content and in the presence of cytoplasmic coats. Immunogold labeling demonstrated that some vesicles within the clusters contained gp58, a cis Golgi marker, and some contained alpha-mannosidase II, a middle/trans Golgi marker in this cell type. Neither marker was detected in the rough ER by immunogold or immunofluorescence labeling. When AlF4- was added during BFA treatment some of the vesicles in the clusters appeared coated. When microsomes were subfractionated into Golgi (light) and rough ER (heavy) fractions on sucrose density gradients, greater than 65% of alpha-mannosidase II and galactosyltransferase activities were found in light fractions (1.14-1.16 g/ml) in both control and BFA-treated lobules. In both cases equally low enzyme activity was recovered in heavier fractions (1.2-1.23 g/ml) containing RNA and alpha-glucosidase activity. However, 5 to 8% of the total recovered RNA consistently codistributed with the Golgi enzyme peak. These results indicate that BFA rapidly inhibits secretion and causes dismantling of the Golgi stacks in pancreatic acinar cells, but clusters of vesicles consisting of bona fide Golgi remnants persist even with prolonged exposure to BFA. Many of the vesicles contain Golgi markers by immunolabeling. By cell fractionation Golgi membrane enzyme activities are recovered in equal amounts in light (Golgi) fractions in both controls and BFA-treated specimens. These findings indicate that in the exocrine pancreas there is a dissociation of BFA's effects on the exocytic pathway: there is a block in transport and Golgi organization is disrupted, but remnant Golgi vesicles and tubules persist and retain Golgi membrane antigens and enzyme activities.     

Benzyloxybenzaldehyde analogues as novel adenylyl cyclase activators.

Several benzyloxybenzaldehyde analogues were prepared and found to have significant inhibitory activity toward neutrophil superoxide formation. Consequently, these compounds were evaluated for cAMP-elevating capability. Among them, benzyloxybenzaldehyde (7), exhibiting activity equivalent to forskolin, was determined as an adenylyl cyclase activator since it elevates cAMP levels by activation of adenylyl cyclase but not by inhibition of phosphodiesterase. Having a chemical structure very different from known adenylyl cyclase activators, compound 7 is recommended by us for use as a new lead compound in the future development of adenylyl cyclase activators.     

Regulation of the adenylyl cyclase signaling system in various types of cultured endothelial cells

We studied the effects of modulators of the adenylyl cyclase pathway on the accumulation of cAMP in endothelial cells isolated from bovine aortas, pig pulmonary arteries, human umbilical veins, and human subcutaneous adipose microvessels. In addition to quantitative differences in the basal levels, cAMP stimulation in different endothelial cell types varied in sensitivity and magnitude in response to both the direct adenylyl cyclase activator forskolin and the β-adrenergic receptor agonist isoproterenol. Furthermore, the ubiquitous phosphodiesterase inhibitor IBMX differentially enhanced both the basal and the stimulated cAMP levels in the various cell types. Histamine caused an elevation of cAMP only in bovine aortic endothelial cells and in human umbilical vein endothelial cells. Treatment of the cells with cholera and pertussis toxins, which uniquely affect G-protein subunits, resulted in divergent elevation of cAMP in the various cells. Thus, in each cell type, a distinct profile of regulation of the cAMP levels was found. Our results suggest that the adenylyl cyclase signaling system in various types of endothelial cells can be differentially regulated at the levels of receptors, G-proteins, adenylyl cyclase, and phosphodiesterase.     

Capacity of the golgi apparatus for biogenesis from the endoplasmic reticulum

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.     

Brefeldin A inhibits protein synthesis in cultured cells.

The fungal metabolite brefeldin A (BFA) is known to disrupt the Golgi apparatus resulting in redistribution of Golgi proteins to the endoplasmic reticulum and inhibition of protein secretion. BFA was found to inhibit protein synthesis in rat glioma C6 cells by up to 70% between 0.1 and 1 microgram/ml. Inhibition was both time-dependent and reversible. BFA inhibited protein synthesis to varying degrees in a number of other cell lines but not in BFA-resistant marsupial kidney cells. The same concentrations of BFA which inhibited protein synthesis, also blocked the inhibitory effects of Pseudomonas exotoxin and ricin on BFA-sensitive cells. BFA, however, was unable to block the inhibition of protein synthesis by the toxins in the resistant marsupial kidney cells.