Golgi Disassembly Research Articles

Molecular Mechanism of Mitotic Golgi Disassembly and Reassembly Revealed by a Defined Reconstitution Assay

In mammalian cells, flat Golgi cisternae closely arrange together to form stacks. During mitosis, the stacked structure undergoes a continuous fragmentation process. The generated mitotic Golgi fragments are distributed into the daughter cells, where they are reassembled into new Golgi stacks. In this study, an in vitro assay has been developed using purified proteins and Golgi membranes to reconstitute the Golgi disassembly and reassembly processes. This technique provides a useful tool to delineate the mechanisms underlying the morphological change. There are two processes during Golgi disassembly: unstacking and vesiculation. Unstacking is mediated by two mitotic kinases, cdc2 and plk, which phosphorylate the Golgi stacking protein GRASP65 and thus disrupt the oligomer of this protein. Vesiculation is mediated by the COPI budding machinery ARF1 and the coatomer complex. When treated with a combination of purified kinases, ARF1 and coatomer, the Golgi membranes were completely fragmented into vesicles. After mitosis, there are also two processes in Golgi reassembly: formation of single cisternae by membrane fusion, and restacking. Cisternal membrane fusion requires two AAA ATPases, p97 and NSF (N-ethylmaleimide-sensitive fusion protein), each of which functions together with specific adaptor proteins. Restacking of the newly formed Golgi cisternae requires dephosphorylation of Golgi stacking proteins by the protein phosphatase PP2A. This systematic study revealed the minimal machinery that controls the mitotic Golgi disassembly and reassembly processes.

AMP-activated Protein Kinase Phosphorylates Golgi-specific Brefeldin A Resistance Factor 1 at Thr1337 to Induce Disassembly of Golgi Apparatus

Sufficiency and depletion of nutrients regulate the cellular activities through the protein phosphorylation reaction; however, many protein substrates remain to be clarified. GBF1 (Golgi-specific brefeldin A resistance factor 1), a guanine nucleotide exchange factor for the ADP-ribosylation factor family associated with the Golgi apparatus, was isolated as a phosphoprotein from the glucose-depleted cells by using the phospho-Akt-substrate antibody, which recognizes the substrate proteins of several protein kinases. The phosphorylation of GBF1 was induced by 2-deoxyglucose (2-DG), which blocks glucose utilization and increases the intracellular AMP concentration, and by AICAR, an AMP-activated protein kinase (AMPK) activator. This phosphorylation was observed in the cells expressing the constitutively active AMPK. The 2-DG-induced phosphorylation of GBF1 was suppressed by Compound C, an AMPK inhibitor, and by the overexpression of the kinase-negative AMPK. Analysis using the deletion and point mutants identified Thr(1337) as the 2-DG-induced phosphorylation site in GBF1, which is phosphorylated by AMPK in vitro. ATP depletion is known to provoke the Golgi apparatus disassembly. Immunofluorescent microscopic analysis with the Golgi markers indicated that GBF1 associates with the fragmented Golgi apparatus in the cells treated with 2-DG and AICAR. The expression of the kinase-negative AMPK and the GBF1 mutant replacing Thr(1337) by Ala prevented the 2-DG-induced Golgi disassembly. These results indicate that GBF1 is a novel AMPK substrate and that the AMPK-mediated phosphorylation of GBF1 at Thr(1337) has a critical role, presumably by attenuating the function of GBF1, in the disassembly of the Golgi apparatus induced under stress conditions that lower the intracellular ATP concentration.

細胞内の膜オルガネラの創成機構 小胞輸送で結ばれた哺乳動物細胞オルガネラの膜創成機構

Organelles are connected and exchange proteins and lipids each other by vesicular transport, while maintaining their specific morphology and components during interphase. At the onset of mitosis, the balance of vesicular transport between organelles is disturbed, resulting in the drastic change of organelle morphology. To understand the coupling mechanisms between organelle morphology and vesicular transport, we developed the reconstitution systems for the Golgi disassembly, the disruption and reformation of the endoplasmic reticulum (ER), mitotic disruption of ER exit sites, and vesicular transport between ER and Golgi in semi-intact cells using interphase or mitotic cytosol, and elucidated biochemical requirements for the processes. In this paper, we will show the molecular links between organelle morphology and transport, especially focused on the mitotic kinases and its regulation of membrane fusion machineries.

Identification of a Redox-sensitive Cysteine in GCP60 That Regulates Its Interaction with Golgin-160

Golgin-160 is ubiquitously expressed in vertebrates. It localizes to the cytoplasmic side of the Golgi and has a large C-terminal coiled-coil domain. The noncoiled-coil N-terminal head domain contains Golgi targeting information, a cryptic nuclear localization signal, and three caspase cleavage sites. Caspase cleavage of the golgin-160 head domain generates different fragments that can translocate to the nucleus by exposing the nuclear localization signal. We have previously shown that GCP60, a Golgi resident protein, interacts weakly with the golgin-160 head domain but has a strong interaction with one of the caspase-generated golgin-160 fragments (residues 140-311). This preferential interaction increases the Golgi retention of the golgin-160 fragment in cells overexpressing GCP60. Here we studied the interaction of golgin-160-(140-311) with GCP60 and identified a single cysteine residue in GCP60 (Cys-463) that is critical for the interaction of the two proteins. Mutation of the cysteine blocked the interaction in vitro and disrupted the ability to retain the golgin-160 fragment at the Golgi in cells. We also found that Cys-463 is redox-sensitive; in its reduced form, interaction with golgin-160 was diminished or abolished, whereas oxidation of the Cys-463 by hydrogen peroxide restored the interaction. In addition, incubation with a nitric oxide donor promoted this interaction in vitro. These findings suggest that nuclear translocation of golgin-160-(140-311) is a highly coordinated event regulated not only by cleavage of the golgin-160 head but also by the oxidation state of GCP60.

Shuttling of G Protein Subunits between the Plasma Membrane and Intracellular Membranes

Heterotrimeric G proteins (alphabetagamma) mediate the majority of signaling pathways in mammalian cells. It is long held that G protein function is localized to the plasma membrane. Here we examined the spatiotemporal dynamics of G protein localization using fluorescence recovery after photobleaching, fluorescence loss in photobleaching, and a photoswitchable fluorescent protein, Dronpa. Unexpectedly, G protein subunits shuttle rapidly (t1/2 < 1 min) between the plasma membrane and intracellular membranes. We show that consistent with such shuttling, G proteins constitutively reside in endomembranes. Furthermore, we show that shuttling is inhibited by 2-bromopalmitate. Thus, contrary to present thought, G proteins do not reside permanently on the plasma membrane but are constantly testing the cytoplasmic surfaces of the plasma membrane and endomembranes to maintain G protein pools in intracellular membranes to establish direct communication between receptors and endomembranes.

Mammalian Sec16/p250 Plays a Role in Membrane Traffic from the Endoplasmic Reticulum

Coat protein complex II (COPII)-coated vesicles/carriers, which mediate export of proteins from the endoplasmic reticulum (ER), are formed at special ER subdomains in mammals, termed ER exit sites or transitional ER. The COPII coat consists of a small GTPase, Sar1, and two protein complexes, Sec23-Sec24 and Sec13-Sec31. Sec23-Sec24 and Sec13-Sec31 appear to constitute the inner and the outermost layers of the COPII coat, respectively. We previously isolated two mammalian proteins (p125 and p250) that bind to Sec23. p125 was found to be a mammalian-specific, phospholipase A(1)-like protein that participates in the organization of ER exit sites. Here we show that p250 is encoded by the KIAA0310 clone and has sequence similarity to yeast Sec16 protein. Although KIAA0310p was found to be localized at ER exit sites, subcellular fractionation revealed its predominant presence in the cytosol. Cytosolic KIAA0310p was recruited to ER membranes in a manner dependent on Sar1. Depletion of KIAA0310p mildly caused disorganization of ER exit sites and delayed protein transport from the ER, suggesting its implication in membrane traffic out of the ER. Overexpression of KIAA0310p affected ER exit sites in a manner different from that of p125. Binding experiments suggested that KIAA0310p interacts with both the inner and the outermost layer coat complexes, whereas p125 binds principally to the inner layer complex. Our results suggest that KIAA0310p, a mammalian homologue of yeast Sec16, builds up ER exit sites in cooperation with p125 and plays a role in membrane traffic from the ER.

Golgi complex disassembly caused by light-activated Calphostin C involves MAPK and PKA

We examined the participation of MAPK and PKA in the Golgi complex disassembly caused by light-activated Calphostin C in HT-29 cells. When these cells were incubated with Calphostin C, fragmentation and dispersal of the Golgi complex was observed as assessed by immunofluorescence microscopy. Electron microscopy analysis showed that clusters of vesicles and large tubule-vesicular membrane structures, resembling the Golgi remnants present in mitotic cells, substituted the Golgi stacks. In addition, Calphostin C treatment caused inhibition of the endocytic route. We confirmed that the Golgi disassembly was not due to PKC inhibition, and suggested, based on the use of specific inhibitors, that other kinases are involved. It was shown that pretreatment with PD98059 and H-89, both inhibitors of MAPK and PKA, respectively, prior to incubation with Calphostin C, caused blockade of the Golgi disassembly, as well as the inhibition of the endocytic pathway caused by this drug. This finding supports the existence of a novel mechanism by which MAPK and PKA may regulate the Golgi breakdown caused by Calphostin C in HT-29 cells.

Mitogen-activated Protein Kinase Kinase 1-dependent Golgi Unlinking Occurs in G2Phase and Promotes the G2/M Cell Cycle Transition

Two controversies have emerged regarding the signaling pathways that regulate Golgi disassembly at the G(2)/M cell cycle transition. The first controversy concerns the role of mitogen-activated protein kinase activator mitogen-activated protein kinase kinase (MEK)1, and the second controversy concerns the participation of Golgi structure in a novel cell cycle "checkpoint." A potential simultaneous resolution is suggested by the hypothesis that MEK1 triggers Golgi unlinking in late G(2) to control G(2)/M kinetics. Here, we show that inhibition of MEK1 by RNA interference or by using the MEK1/2-specific inhibitor U0126 delayed the passage of synchronized HeLa cells into M phase. The MEK1 requirement for normal mitotic entry was abrogated if Golgi proteins were dispersed before M phase by treatment of cells with brefeldin A or if GRASP65, which links Golgi stacks into a ribbon network, was depleted. Imaging revealed that unlinking of the Golgi apparatus begins before M phase, is independent of cyclin-dependent kinase 1 activation, and requires MEK signaling. Furthermore, expression of the GRASP family member GRASP55 after alanine substitution of its MEK1-dependent mitotic phosphorylation sites inhibited both late G(2) Golgi unlinking and the G(2)/M transition. Thus, MEK1 plays an in vivo role in Golgi reorganization, which regulates cell cycle progression.

Nordihydroguaiaretic Acid Affects Multiple Dynein-Dynactin Functions in Interphase and Mitotic Cells

Nordihydroguaiaretic acid (NDGA), a well known lipoxygenase inhibitor, actually has pleiotropic effects on cells, which include cell proliferation, apoptosis, differentiation, and chemotaxis. We and others have shown previously that this compound causes Golgi disassembly by an unknown mechanism. In this study, we show that, in parallel with Golgi disassembly, NDGA induces the accumulation of the microtubule minus-end-directed motor dynein-dynactin complex at the centrosome, where microtubules minus-ends lie. Concomitant with this accumulation, dynein-dynactin-interacting proteins, such as ZW10 and EB1, were also redistributed to the centrosomal region. In cells where microtubules were depolymerized by nocodazole, NDGA promoted the formation of filaments consisting of dynein-dynactin and its interacting proteins, suggesting that it stimulates the association of these proteins in an ordered, not random, manner. Loss of dynactin function abolished not only NDGA-induced redistribution in intact cells but also filament formation in nocodazole-treated cells. The latter finding implies that dynactin is a key molecule for the association between dynein-dynactin and its interacting proteins. In mitotic cells, NDGA induced robust accumulation of dyneindynactin and its interacting proteins at the spindle poles. These results taken together suggest that NDGA perturbs membrane traffic by affecting the function of the microtubule motor dynein-dynactin complex and its auxiliary proteins. To our knowledge, NDGA is the first case of a reagent that can modulate dynein-dynactin-related processes.

GCP60 Preferentially Interacts with a Caspase-generated Golgin-160 Fragment

Golgin-160, a ubiquitous protein in vertebrates, localizes to the cytoplasmic face of the Golgi complex. Golgin-160 has a large coiled-coil C-terminal domain and a non-coiled-coil N-terminal ("head") domain. The head domain contains important motifs, including a nuclear localization signal, a Golgi targeting domain, and three aspartates that are recognized by caspases during apoptosis. Some of the caspase cleavage products accumulate in the nucleus when overexpressed. Expression of a non-cleavable form of golgin-160 impairs apoptosis induced by some pro-apoptotic stimuli; thus cleavage of golgin-160 appears to play a role in apoptotic signaling. We used a yeast two-hybrid assay to screen for interactors of the golgin-160 head and identified GCP60 (Golgi complex-associated protein of 60 kDa). Further analysis demonstrated that GCP60 interacts preferentially with one of the golgin-160 caspase cleavage fragments (residues 140-311). This strong interaction prevented the golgin-160 fragment from accumulating in the nucleus when this fragment and GCP60 were overexpressed. In addition, cells overexpressing GCP60 were more sensitive to apoptosis induced by staurosporine, suggesting that nuclear-localized golgin-160-(140-311) might promote cell survival. Our results suggest a potential mechanism for regulating the nuclear translocation and potential functions of golgin-160 fragments.

Isoform-specific Interaction of Golgin-160 with the Golgi-associated Protein PIST

Golgin-160 belongs to the golgin family of Golgi-localized proteins, which have been implicated in Golgi structure and function. Golgin-160 possesses an N-terminal non-coiled-coil "head" domain followed by an extensive coiled-coil region. Using the N-terminal head domain of golgin-160 as bait in a yeast two-hybrid screen, the postsynaptic density-95/Discs large/zona occludens-1 (PDZ) domain protein interacting specifically with TC10 (PIST) was identified to interact with golgin-160. PIST (also known as GOPC, CAL, and FIG) has been implicated in the trafficking of a subset of plasma membrane proteins, supporting a role of golgin-160 in vesicular trafficking. Golgin-160 and PIST colocalize to Golgi membranes and interact in vivo. Glutathione S-transferase binding experiments identified an internal region of PIST that includes a coiled-coil domain, which interacts directly with golgin-160. Similar binding experiments identified a leucine-rich repeat within golgin-160 necessary for interaction with PIST. Therefore, our data suggest that golgin-160 may participate in PIST-dependent trafficking of cargo. Interestingly, we also discovered a widely expressed isoform of golgin-160, golgin-160B, which lacks the exon encoding the leucine repeat that mediates binding to PIST. As predicted, golgin-160B was unable to bind PIST. Full-length golgin-160 and golgin-160B may link distinct subsets of proteins to effect specific membrane trafficking pathways.

Protein Phosphatase 2A Activity Associated with Golgi Membranes during the G2/M Phase May Regulate Phosphorylation of ERK2

The extracellular signal-regulated kinase (ERK) 1 and 2 proteins are mitogen-activated protein kinase (MAPK) members that regulate cell proliferation and differentiation. ERK proteins are activated exclusively by MAPK kinase 1 and 2 phosphorylation of threonine and tyrosine residues located within the conserved TXY MAPK activation motif. Although dual phosphorylation of Thr and Tyr residues confers full activation of ERK, in vitro studies suggest that a single phosphorylation on either Thr or Tyr may yield partial ERK activity. Previously, we have demonstrated that phosphorylation of the tyrosine residue (Tyr(P) ERK) may be involved in regulating the Golgi complex structure during the G2 and M phases of the cell cycle (Cha, H., and Shapiro, P. (2001) J. Cell Biol. 153, 1355-1368). In the present study, we examined mechanisms for generating Tyr(P) ERK by determining cell cycle-dependent changes in localized phosphatase activity. Using fractionated nuclei-free cell lysates, we find increased serine/threonine phosphatase activity associated with Golgi-enriched membranes in cells synchronized in the late G2/early M phase as compared with G1 phase cells. The addition of phosphatase inhibitors in combination with immunodepletion assays identified this activity to be related to protein phosphatase 2A (PP2A). The increased activity was accounted for by elevated PP2A association with mitotic Golgi membranes as well as increased catalytic activity after normalization of PP2A protein levels in the phosphatase assays. These data indicate that localized changes in PP2A activity may be involved in regulating proteins involved in Golgi disassembly as cells enter mitosis.

Norwalk virus N-terminal nonstructural protein is associated with disassembly of the Golgi complex in transfected cells.

Norwalk virus is the prototype strain for members of the genus Norovirus in the family Caliciviridae, which are associated with epidemic gastroenteritis in humans. The nonstructural protein encoded in the N-terminal region of the first open reading frame (ORF1) of the Norwalk virus genome is analogous in gene order to proteins 2A and 2B of the picornaviruses; the latter is known for its membrane-associated activities. Confocal microscopy imaging of cells transfected with a vector plasmid that provided expression of the entire Norwalk virus N-terminal protein (amino acids 1 to 398 of the ORF1 polyprotein) showed colocalization of this protein with cellular proteins of the Golgi apparatus. Furthermore, this colocalization was characteristically associated with a visible disassembly of the Golgi complex into discrete aggregates. Deletion of a predicted hydrophobic region (amino acids 360 to 379) in a potential 2B-like (2BL) region (amino acids 301 to 398) near the C terminus of the Norwalk virus N-terminal protein reduced Golgi colocalization and disassembly. Confocal imaging was conducted to examine the expression characteristics of fusion proteins in which the 2BL region from the N-terminal protein of Norwalk virus (a genogroup I norovirus) or MD145 (a genogroup II norovirus) was fused to the C terminus of enhanced green fluorescent protein. Expression of each fusion protein in cells showed evidence for its colocalization with the Golgi apparatus. These data indicate that the N-terminal protein of Norwalk virus interacts with the Golgi apparatus and may play a 2BL role in the induction of intracellular membrane rearrangements associated with positive-strand RNA virus replication in cells.

Dispersal of Golgi matrix proteins during mitotic Golgi disassembly.

During mitosis, the mammalian Golgi disassembles into numerous vesicles and larger membrane structures referred to as clusters or remnants. Following mitosis, the vesicles and clusters reassemble to form an intact Golgi in each daughter cell. One model of Golgi biogenesis states that Golgi matrix proteins remain assembled in mitotic clusters and then serve as a template for Golgi reassembly. To test this idea, we performed a 3D-computational analysis of mitotic cells to determine the extent to which these proteins remain in mitotic clusters. As a control we used brefeldin A-induced Golgi disassembly which causes dispersal of Golgi enzymes, but leaves matrix proteins in remnant structures. Unlike brefeldin A-treated cells, in which matrix proteins were clearly sorted from non-matrix proteins, we observed extensive dispersal of matrix proteins in metaphase cells with no evidence of differential sorting of these proteins from other Golgi proteins. The extensive disassembly of matrix proteins argues against their participation in a stable template and supports a self-assembly mode of Golgi biogenesis.

A role for Arf1 in mitotic Golgi disassembly, chromosome segregation, and cytokinesis.

In mitosis, chromosome, cytoskeleton, and organelle dynamics must be coordinated for successful cell division. Here, we present evidence for a role for Arf1, a small GTPase associated with the Golgi apparatus, in the orchestration of mitotic Golgi breakdown, chromosome segregation, and cytokinesis. We show that early in mitosis Arf1 becomes inactive and dissociates from Golgi membranes. This is followed by the dispersal of numerous Arf1-dependent peripheral Golgi proteins and subsequent Golgi disassembly. If Arf1 is kept in an active state by treatment with the small molecule H89 or expression of its GTP-locked form, intact Golgi membranes with bound peripheral proteins persist throughout mitosis. These cells enter mitosis but exhibit gross defects in chromosome segregation and cytokinetic furrow ingression. These findings suggest that mitotic Golgi disassembly depends on Arf1 inactivation and is used by the cell to disperse numerous peripheral Golgi proteins for coordinating the behavior of Golgi membranes, chromosomes, and cytoskeleton during mitosis.

The localization and phosphorylation of p47 are important for Golgi disassembly-assembly during the cell cycle.

In mammalian cells, the Golgi apparatus is disassembled at the onset of mitosis and reassembled at the end of mitosis. This disassembly–reassembly is generally believed to be essential for the equal partitioning of Golgi into two daughter cells. For Golgi disassembly, membrane fusion, which is mediated by NSF and p97, needs to be blocked. For the NSF pathway, the tethering of p115-GM130 is disrupted by the mitotic phosphorylation of GM130, resulting in the inhibition of NSF-mediated fusion. In contrast, the p97/p47 pathway does not require p115-GM130 tethering, and its mitotic inhibitory mechanism has been unclear. Now, we have found that p47, which mainly localizes to the nucleus during interphase, is phosphorylated on Serine-140 by Cdc2 at mitosis. The phosphorylated p47 does not bind to Golgi membranes. An in vitro assay shows that this phosphorylation is required for Golgi disassembly. Microinjection of p47(S140A), which is unable to be phosphorylated, allows the cell to keep Golgi stacks during mitosis and has no effect on the equal partitioning of Golgi into two daughter cells, suggesting that Golgi fragmentation-dispersion may not be obligatory for equal partitioning even in mammalian cells.

Golgi disassembly in apoptosis: cause or effect?

The Golgi complex undergoes a dramatic disassembly process during apoptosis. Some Golgi proteins implicated in Golgi structure and vesicle transport are cleaved during apoptosis, and expression of noncleavable mutants of these proteins delays Golgi disassembly after pro-apoptotic stimuli. Cleavage of Golgi structural proteins and subsequent disassembly of the organelle could simply be the result of the apoptotic process. However, recent studies raise the intriguing possibility that cleavage of Golgi proteins during apoptosis might be required for more than disassembly of the organelle.

Differential Mobilization of Newly Synthesized Cholesterol and Biosynthetic Sterol Precursors from Cells

Previous work demonstrates that the biosynthetic precursor of cholesterol, desmosterol, is released from cells and that its efflux to high density lipoprotein or phosphatidylcholine vesicles is greater than that of newly synthesized cholesterol (Johnson, W. J., Fischer, R. T., Phillips, M. C., and Rothblat, G. H. (1995) J. Biol. Chem. 270, 25037-25046). Here we report that the release of individual precursor sterols varies with the efflux of newly synthesized zymosterol being greater than that of lathosterol and both exceeding that of newly synthesized cholesterol when using either methyl-beta-cyclodextrin or complete serum as acceptors. The transfer of newly synthesized lathosterol to methyl-beta-cyclodextrin was inhibited by actin polymerization but not by Golgi disassembly whereas that of newly synthesized cholesterol was inhibited by both conditions. Newly synthesized lathosterol associated with cellular detergent-resistant membranes more rapidly than newly synthesized cholesterol. Upon efflux to serum, newly synthesized cholesterol precursors associated with both high and low density lipoproteins. Stimulation of the formation of direct endoplasmic reticulum-plasma membrane contacts was accompanied by enhanced efflux of newly synthesized lathosterol but not of newly synthesized cholesterol to serum acceptors. The data indicate that the efflux of cholesterol precursors differs not only from that of cholesterol but also from each other, with the more polar zymosterol being more avidly effluxed. Moreover, the results suggest that the intracellular routing of cholesterol precursors differs from that of newly synthesized cholesterol and implicates a potential role for the actin cytoskeleton and endoplasmic reticulum-plasma membrane contacts in the efflux of lathosterol.

Golgi apparatus partitioning during cell division.

This review discusses the mitotic segregation of the Golgi apparatus. The results from classical biochemical and morphological studies have suggested that in mammalian cells this organelle remains distinct during mitosis, although highly fragmented through the formation of mitotic Golgi clusters of small tubules and vesicles. Shedding of free Golgi-derived vesicles would consume Golgi clusters and disperse this organelle throughout the cytoplasm. Vesicles could be partitioned in a stochastic and passive way between the two daughter cells and act as a template for the reassembly of this key organelle. This model has recently been modified by results obtained using GFP- or HRP-tagged Golgi resident enzymes, live cell imaging and electron microscopy. Results obtained with these techniques show that the mitotic Golgi clusters are stable entities throughout mitosis that partition in a microtubule spindle-dependent fashion. Furthermore, a newer model proposes that at the onset of mitosis, the Golgi apparatus completely loses its identity and is reabsorbed into the endoplasmic reticulum. This suggests that the partitioning of the Golgi apparatus is entirely dependent on the partitioning of the endoplasmic reticulum. We critically discuss both models and summarize what is known about the molecular mechanisms underlying the Golgi disassembly and reassembly during and after mitosis. We will also review how the study of the Golgi apparatus during mitosis in other organisms can answer current questions and perhaps reveal novel mechanisms.

Inactivation of Galpha(z) causes disassembly of the Golgi apparatus.

We showed previously that overexpression of the alpha subunit of G(z) or G(i2) suppresses nordihydroguaiaretic acid-induced Golgi disassembly. To determine whether the active form of Galpha is required to maintain the structure of the Golgi apparatus, we examined the effects of a series of Galpha GAPs, regulators of G protein signaling (RGS) proteins, on the Golgi structure. Expression of RGSZ1 or RGSZ2, both of which exhibit high selectivity for Galpha(z), markedly induced dispersal of the Golgi apparatus, whereas expression of RGS proteins that are rather selective for Galpha(q) or other Galpha(i) species did not. A mutated RGSZ1, which is deficient in the interaction with Galpha(z), did not induce Golgi disassembly. These results suggest that the active form of Galpha(z), but not Galpha(i2), is crucial for maintenance of the structure of the Golgi apparatus. Consistent with this idea, Golgi disruption also took place in cells transfected with a dominant-negative Galpha(z) mutant. Although previous studies showed that the expression of Galpha(z) is confined to neuronal cells and platelets, immunofluorescence and mRNA expression analyses revealed that it is also expressed, albeit at low levels, in non-neuronal cells, and is located in the Golgi apparatus. These results taken together suggest a general regulatory role for Galpha(z) in the control of the Golgi structure.