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The coiled-coil membrane protein golgin-84 is a novel rab effector required for Golgi ribbon formation.

Diao A, Rahman D, Pappin DJ, Lucocq J, Lowe M - J. Cell Biol. (2003)

Bottom Line: Using cryoelectron microscopy we could localize golgin-84 to the cis-Golgi network and found that it is enriched on tubules emanating from the lateral edges of, and often connecting, Golgi stacks.These mini-stacks are able to carry out protein transport, though with reduced efficiency compared with a normal Golgi apparatus.Our results suggest that golgin-84 plays a key role in the assembly and maintenance of the Golgi ribbon in mammalian cells.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, University of Manchester, Manchester M13 9PT, UK.

ABSTRACT
Fragmentation of the mammalian Golgi apparatus during mitosis requires the phosphorylation of a specific subset of Golgi-associated proteins. We have used a biochemical approach to characterize these proteins and report here the identification of golgin-84 as a novel mitotic target. Using cryoelectron microscopy we could localize golgin-84 to the cis-Golgi network and found that it is enriched on tubules emanating from the lateral edges of, and often connecting, Golgi stacks. Golgin-84 binds to active rab1 but not cis-Golgi matrix proteins. Overexpression or depletion of golgin-84 results in fragmentation of the Golgi ribbon. Strikingly, the Golgi ribbon is converted into mini-stacks constituting only approximately 25% of the volume of a normal Golgi apparatus upon golgin-84 depletion. These mini-stacks are able to carry out protein transport, though with reduced efficiency compared with a normal Golgi apparatus. Our results suggest that golgin-84 plays a key role in the assembly and maintenance of the Golgi ribbon in mammalian cells.

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Golgin-84 depletion results in a significant loss of membrane from the Golgi apparatus and an exaggerated ER. Control (a) or golgin-84 RNAi–treated HeLa cells (b and c) were fixed and embedded in Epon for electron microscopy. Large arrows indicate Golgi membranes whereas small arrows indicate the ER. Note the decrease in size of the Golgi apparatus and the exaggerated and swollen ER in golgin-84–depleted cells. (d) Quantitation of the amount of membrane in Golgi cisternae, Golgi vesicles, ER, and nuclear envelope (NE) in golgin-84–depleted cells relative to control cells expressed as density of membrane surface in cell volume. For controls and RNAi samples, n = 23 and 22 micrographs, respectively. Bars, 200 nm.
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fig7: Golgin-84 depletion results in a significant loss of membrane from the Golgi apparatus and an exaggerated ER. Control (a) or golgin-84 RNAi–treated HeLa cells (b and c) were fixed and embedded in Epon for electron microscopy. Large arrows indicate Golgi membranes whereas small arrows indicate the ER. Note the decrease in size of the Golgi apparatus and the exaggerated and swollen ER in golgin-84–depleted cells. (d) Quantitation of the amount of membrane in Golgi cisternae, Golgi vesicles, ER, and nuclear envelope (NE) in golgin-84–depleted cells relative to control cells expressed as density of membrane surface in cell volume. For controls and RNAi samples, n = 23 and 22 micrographs, respectively. Bars, 200 nm.

Mentions: To characterize the effects of golgin-84 depletion upon Golgi morphology in more detail, siRNA-treated cells were analyzed by electron microscopy. Interestingly, the Golgi apparatus appeared significantly smaller in golgin-84–depleted cells compared with control cells (Fig. 7, a and b). Quantitation revealed that ∼75% of Golgi cisternae and ∼70% of Golgi tubulo-vesicular membranes were lost upon golgin-84 depletion (Fig. 7 d). The overall morphology was not, however, dramatically affected. The number of stacked cisternae per stack and the ratio of membrane in stacked cisternae versus tubulo-vesicular profiles were similar in golgin-84–depleted cells and control cells (unpublished data). Depletion of golgin-84 also had a marked effect upon the ER, which was grossly exaggerated compared with control cells, forming an elaborated network extending throughout the cytoplasm (Fig. 7 c). The lumen of the ER often appeared swollen with electron-dense material, suggesting that an accumulation of proteins had occurred there. Quantitation revealed that the ER was ∼1.7 times larger in golgin-84–depleted cells compared with control cells, while the nuclear envelope was only marginally affected. Thus, depletion of golgin-84 results in a dramatic loss of Golgi membranes and a corresponding increase in the size of the ER.


The coiled-coil membrane protein golgin-84 is a novel rab effector required for Golgi ribbon formation.

Diao A, Rahman D, Pappin DJ, Lucocq J, Lowe M - J. Cell Biol. (2003)

Golgin-84 depletion results in a significant loss of membrane from the Golgi apparatus and an exaggerated ER. Control (a) or golgin-84 RNAi–treated HeLa cells (b and c) were fixed and embedded in Epon for electron microscopy. Large arrows indicate Golgi membranes whereas small arrows indicate the ER. Note the decrease in size of the Golgi apparatus and the exaggerated and swollen ER in golgin-84–depleted cells. (d) Quantitation of the amount of membrane in Golgi cisternae, Golgi vesicles, ER, and nuclear envelope (NE) in golgin-84–depleted cells relative to control cells expressed as density of membrane surface in cell volume. For controls and RNAi samples, n = 23 and 22 micrographs, respectively. Bars, 200 nm.
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Related In: Results  -  Collection

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fig7: Golgin-84 depletion results in a significant loss of membrane from the Golgi apparatus and an exaggerated ER. Control (a) or golgin-84 RNAi–treated HeLa cells (b and c) were fixed and embedded in Epon for electron microscopy. Large arrows indicate Golgi membranes whereas small arrows indicate the ER. Note the decrease in size of the Golgi apparatus and the exaggerated and swollen ER in golgin-84–depleted cells. (d) Quantitation of the amount of membrane in Golgi cisternae, Golgi vesicles, ER, and nuclear envelope (NE) in golgin-84–depleted cells relative to control cells expressed as density of membrane surface in cell volume. For controls and RNAi samples, n = 23 and 22 micrographs, respectively. Bars, 200 nm.
Mentions: To characterize the effects of golgin-84 depletion upon Golgi morphology in more detail, siRNA-treated cells were analyzed by electron microscopy. Interestingly, the Golgi apparatus appeared significantly smaller in golgin-84–depleted cells compared with control cells (Fig. 7, a and b). Quantitation revealed that ∼75% of Golgi cisternae and ∼70% of Golgi tubulo-vesicular membranes were lost upon golgin-84 depletion (Fig. 7 d). The overall morphology was not, however, dramatically affected. The number of stacked cisternae per stack and the ratio of membrane in stacked cisternae versus tubulo-vesicular profiles were similar in golgin-84–depleted cells and control cells (unpublished data). Depletion of golgin-84 also had a marked effect upon the ER, which was grossly exaggerated compared with control cells, forming an elaborated network extending throughout the cytoplasm (Fig. 7 c). The lumen of the ER often appeared swollen with electron-dense material, suggesting that an accumulation of proteins had occurred there. Quantitation revealed that the ER was ∼1.7 times larger in golgin-84–depleted cells compared with control cells, while the nuclear envelope was only marginally affected. Thus, depletion of golgin-84 results in a dramatic loss of Golgi membranes and a corresponding increase in the size of the ER.

Bottom Line: Using cryoelectron microscopy we could localize golgin-84 to the cis-Golgi network and found that it is enriched on tubules emanating from the lateral edges of, and often connecting, Golgi stacks.These mini-stacks are able to carry out protein transport, though with reduced efficiency compared with a normal Golgi apparatus.Our results suggest that golgin-84 plays a key role in the assembly and maintenance of the Golgi ribbon in mammalian cells.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, University of Manchester, Manchester M13 9PT, UK.

ABSTRACT
Fragmentation of the mammalian Golgi apparatus during mitosis requires the phosphorylation of a specific subset of Golgi-associated proteins. We have used a biochemical approach to characterize these proteins and report here the identification of golgin-84 as a novel mitotic target. Using cryoelectron microscopy we could localize golgin-84 to the cis-Golgi network and found that it is enriched on tubules emanating from the lateral edges of, and often connecting, Golgi stacks. Golgin-84 binds to active rab1 but not cis-Golgi matrix proteins. Overexpression or depletion of golgin-84 results in fragmentation of the Golgi ribbon. Strikingly, the Golgi ribbon is converted into mini-stacks constituting only approximately 25% of the volume of a normal Golgi apparatus upon golgin-84 depletion. These mini-stacks are able to carry out protein transport, though with reduced efficiency compared with a normal Golgi apparatus. Our results suggest that golgin-84 plays a key role in the assembly and maintenance of the Golgi ribbon in mammalian cells.

Show MeSH
Related in: MedlinePlus