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Peri-Golgi vesicles contain retrograde but not anterograde proteins consistent with the cisternal progression model of intra-Golgi transport.

Martinez-Menárguez JA, Prekeris R, Oorschot VM, Scheller R, Slot JW, Geuze HJ, Klumperman J - J. Cell Biol. (2001)

Bottom Line: A cisternal progression mode of intra-Golgi transport requires that Golgi resident proteins recycle by peri-Golgi vesicles, whereas the alternative model of vesicular transport predicts anterograde cargo proteins to be present in such vesicles.We found significant levels of the Golgi resident enzyme mannosidase II and the transport machinery proteins giantin, KDEL-receptor, and rBet1 in coatomer protein I-coated cisternal rims and peri-Golgi vesicles.By contrast, when cells expressed vesicular stomatitis virus protein G this anterograde marker was largely absent from the peri-Golgi vesicles.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, School of Medicine, University of Murcia, 30071 Murcia, Spain.

ABSTRACT
A cisternal progression mode of intra-Golgi transport requires that Golgi resident proteins recycle by peri-Golgi vesicles, whereas the alternative model of vesicular transport predicts anterograde cargo proteins to be present in such vesicles. We have used quantitative immuno-EM on NRK cells to distinguish peri-Golgi vesicles from other vesicles in the Golgi region. We found significant levels of the Golgi resident enzyme mannosidase II and the transport machinery proteins giantin, KDEL-receptor, and rBet1 in coatomer protein I-coated cisternal rims and peri-Golgi vesicles. By contrast, when cells expressed vesicular stomatitis virus protein G this anterograde marker was largely absent from the peri-Golgi vesicles. These data suggest a role of peri-Golgi vesicles in recycling of Golgi residents, rather than an important role in anterograde transport.

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Immunogold labeling of Man II (A, B, and D, 10 nm gold; C, 15 nm gold) on ultrathin cryosections of NRK cells, showing the presence of Man II in the cis-medial Golgi cisternae (G) and in lateral rims of the cisternae and associated vesicular profiles (arrows). Note that the relative distribution of Man II between cisternae and associated vesicles varies considerably between Golgi's. The arrowhead in A points to a Man II–positive Golgi lateral rim with a clearly visible COP coat. The arrow in C points to a Man II–positive vesicle with a visible COP coat. Bars, 200 nm.
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fig1: Immunogold labeling of Man II (A, B, and D, 10 nm gold; C, 15 nm gold) on ultrathin cryosections of NRK cells, showing the presence of Man II in the cis-medial Golgi cisternae (G) and in lateral rims of the cisternae and associated vesicular profiles (arrows). Note that the relative distribution of Man II between cisternae and associated vesicles varies considerably between Golgi's. The arrowhead in A points to a Man II–positive Golgi lateral rim with a clearly visible COP coat. The arrow in C points to a Man II–positive vesicle with a visible COP coat. Bars, 200 nm.

Mentions: Man II is a prototypical Golgi resident enzyme with a short transmembrane domain and a luminal catalytic domain involved in N-linked glycosylation (Moremen et al., 1991). To select a cell type yielding high Man II labeling in IEM, we performed immunogold labeling for Man II in various tissues (rat pancreatic islets, exocrine pancreas, and liver) and cell-lines (PC12, CHO, HepG2, and NRK). A high overall labeling density combined with a clear cis-to-trans polarity of the Golgi stacks was found in NRK cells, which we therefore chose for further study. The Golgi complex of NRK cells typically consisted of five cisternae (G1–G5 from cis to trans), although occasionally a larger number of cisternae were observed. In a previous study on NRK cells using the enzymatic immunoperoxidase labeling procedure, Man II was localized to one to three medial cisternae (Velasco et al., 1993). By our immunogold labeling approach, we found the majority of Man II label in G2, lower amounts in G1, G3, and G4, whereas G5 was almost devoid of label (Fig. 1; Table I).


Peri-Golgi vesicles contain retrograde but not anterograde proteins consistent with the cisternal progression model of intra-Golgi transport.

Martinez-Menárguez JA, Prekeris R, Oorschot VM, Scheller R, Slot JW, Geuze HJ, Klumperman J - J. Cell Biol. (2001)

Immunogold labeling of Man II (A, B, and D, 10 nm gold; C, 15 nm gold) on ultrathin cryosections of NRK cells, showing the presence of Man II in the cis-medial Golgi cisternae (G) and in lateral rims of the cisternae and associated vesicular profiles (arrows). Note that the relative distribution of Man II between cisternae and associated vesicles varies considerably between Golgi's. The arrowhead in A points to a Man II–positive Golgi lateral rim with a clearly visible COP coat. The arrow in C points to a Man II–positive vesicle with a visible COP coat. Bars, 200 nm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199342&req=5

fig1: Immunogold labeling of Man II (A, B, and D, 10 nm gold; C, 15 nm gold) on ultrathin cryosections of NRK cells, showing the presence of Man II in the cis-medial Golgi cisternae (G) and in lateral rims of the cisternae and associated vesicular profiles (arrows). Note that the relative distribution of Man II between cisternae and associated vesicles varies considerably between Golgi's. The arrowhead in A points to a Man II–positive Golgi lateral rim with a clearly visible COP coat. The arrow in C points to a Man II–positive vesicle with a visible COP coat. Bars, 200 nm.
Mentions: Man II is a prototypical Golgi resident enzyme with a short transmembrane domain and a luminal catalytic domain involved in N-linked glycosylation (Moremen et al., 1991). To select a cell type yielding high Man II labeling in IEM, we performed immunogold labeling for Man II in various tissues (rat pancreatic islets, exocrine pancreas, and liver) and cell-lines (PC12, CHO, HepG2, and NRK). A high overall labeling density combined with a clear cis-to-trans polarity of the Golgi stacks was found in NRK cells, which we therefore chose for further study. The Golgi complex of NRK cells typically consisted of five cisternae (G1–G5 from cis to trans), although occasionally a larger number of cisternae were observed. In a previous study on NRK cells using the enzymatic immunoperoxidase labeling procedure, Man II was localized to one to three medial cisternae (Velasco et al., 1993). By our immunogold labeling approach, we found the majority of Man II label in G2, lower amounts in G1, G3, and G4, whereas G5 was almost devoid of label (Fig. 1; Table I).

Bottom Line: A cisternal progression mode of intra-Golgi transport requires that Golgi resident proteins recycle by peri-Golgi vesicles, whereas the alternative model of vesicular transport predicts anterograde cargo proteins to be present in such vesicles.We found significant levels of the Golgi resident enzyme mannosidase II and the transport machinery proteins giantin, KDEL-receptor, and rBet1 in coatomer protein I-coated cisternal rims and peri-Golgi vesicles.By contrast, when cells expressed vesicular stomatitis virus protein G this anterograde marker was largely absent from the peri-Golgi vesicles.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, School of Medicine, University of Murcia, 30071 Murcia, Spain.

ABSTRACT
A cisternal progression mode of intra-Golgi transport requires that Golgi resident proteins recycle by peri-Golgi vesicles, whereas the alternative model of vesicular transport predicts anterograde cargo proteins to be present in such vesicles. We have used quantitative immuno-EM on NRK cells to distinguish peri-Golgi vesicles from other vesicles in the Golgi region. We found significant levels of the Golgi resident enzyme mannosidase II and the transport machinery proteins giantin, KDEL-receptor, and rBet1 in coatomer protein I-coated cisternal rims and peri-Golgi vesicles. By contrast, when cells expressed vesicular stomatitis virus protein G this anterograde marker was largely absent from the peri-Golgi vesicles. These data suggest a role of peri-Golgi vesicles in recycling of Golgi residents, rather than an important role in anterograde transport.

Show MeSH
Related in: MedlinePlus