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Involvement of the transmembrane protein p23 in biosynthetic protein transport.

Rojo M, Pepperkok R, Emery G, Kellner R, Stang E, Parton RG, Gruenberg J - J. Cell Biol. (1997)

Bottom Line: Moreover, we find that p23 cytoplasmic domain is not involved in COP I membrane recruitment.Our data demonstrate that microinjected antibodies against the cytoplasmic tail of p23 inhibit G protein transport from the cis-Golgi network/ intermediate compartment to the cell surface, suggesting that p23 function is required for the transport of transmembrane cargo molecules.These observations together with the fact that p23 is a highly abundant component in the intermediate compartment, lead us to propose that p23 contributes to membrane structure, and that this contribution is necessary for efficient segregation and transport.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Geneva, 1211 Geneva 4, Switzerland.

ABSTRACT
Here, we report the localization and characterization of BHKp23, a member of the p24 family of transmembrane proteins, in mammalian cells. We find that p23 is a major component of tubulovesicular membranes at the cis side of the Golgi complex (estimated density: 12,500 copies/micron2 membrane surface area, or approximately 30% of the total protein). Our data indicate that BHKp23-containing membranes are part of the cis-Golgi network/intermediate compartment. Using the G protein of vesicular stomatitis virus as a transmembrane cargo molecule, we find that p23 membranes are an obligatory station in forward biosynthetic membrane transport, but that p23 itself is absent from transport vesicles that carry the G protein to and beyond the Golgi complex. Our data show that p23 is not present to any significant extent in coat protein (COP) I-coated vesicles generated in vitro and does not colocalize with COP I buds and vesicles. Moreover, we find that p23 cytoplasmic domain is not involved in COP I membrane recruitment. Our data demonstrate that microinjected antibodies against the cytoplasmic tail of p23 inhibit G protein transport from the cis-Golgi network/ intermediate compartment to the cell surface, suggesting that p23 function is required for the transport of transmembrane cargo molecules. These observations together with the fact that p23 is a highly abundant component in the intermediate compartment, lead us to propose that p23 contributes to membrane structure, and that this contribution is necessary for efficient segregation and transport.

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COP I and p23 distribution. The  indicated cell types were fixed and processed for immunofluorescence, or treated  with 10 μM nocodazole for 2 h before fixation (+ nocodazole). Images were processed and merged (overlay) as in Fig. 3.  The inset shows a higher magnification of  the area indicated by an arrowhead.  Whereas both COP I and p23 localized to  the perinuclear Golgi region in all cell  types, the distribution of the two proteins  was clearly different even in the absence  of nocodazole. It is not clear to what extent overlapping signal (yellow) is due to  colocalization of the proteins or to superimposition of two close but separated fluorescent signals. Bar, 5 μm.
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Figure 8: COP I and p23 distribution. The indicated cell types were fixed and processed for immunofluorescence, or treated with 10 μM nocodazole for 2 h before fixation (+ nocodazole). Images were processed and merged (overlay) as in Fig. 3. The inset shows a higher magnification of the area indicated by an arrowhead. Whereas both COP I and p23 localized to the perinuclear Golgi region in all cell types, the distribution of the two proteins was clearly different even in the absence of nocodazole. It is not clear to what extent overlapping signal (yellow) is due to colocalization of the proteins or to superimposition of two close but separated fluorescent signals. Bar, 5 μm.

Mentions: Electron microscopy studies have localized COP I to the Golgi complex and to the transitional area between ER and Golgi (Oprins et al., 1993; Griffiths et al., 1995). Moreover, Sohn et al. (1996) have reported that rabbit p23 is enriched in COP I–coated vesicles and that p23 cytoplasmic tail peptides have the ability to recruit COP I, leading to the proposal that p23 represents a Golgi-specific receptor for COP I that is involved in the formation of COP I–coated vesicles. We therefore compared the localization of COP I with that of p23. Although both p23 and COP I exhibited a characteristic Golgi-like, perinuclear localization in different cell types, the distribution of the two proteins did not coincide (Fig. 8, CHO, Vero, and HeLa). This differential distribution was also evident after nocodazole-induced formation of dispersed Golgi stacks (Fig. 8, HeLa [+nocodazole]).


Involvement of the transmembrane protein p23 in biosynthetic protein transport.

Rojo M, Pepperkok R, Emery G, Kellner R, Stang E, Parton RG, Gruenberg J - J. Cell Biol. (1997)

COP I and p23 distribution. The  indicated cell types were fixed and processed for immunofluorescence, or treated  with 10 μM nocodazole for 2 h before fixation (+ nocodazole). Images were processed and merged (overlay) as in Fig. 3.  The inset shows a higher magnification of  the area indicated by an arrowhead.  Whereas both COP I and p23 localized to  the perinuclear Golgi region in all cell  types, the distribution of the two proteins  was clearly different even in the absence  of nocodazole. It is not clear to what extent overlapping signal (yellow) is due to  colocalization of the proteins or to superimposition of two close but separated fluorescent signals. Bar, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2140216&req=5

Figure 8: COP I and p23 distribution. The indicated cell types were fixed and processed for immunofluorescence, or treated with 10 μM nocodazole for 2 h before fixation (+ nocodazole). Images were processed and merged (overlay) as in Fig. 3. The inset shows a higher magnification of the area indicated by an arrowhead. Whereas both COP I and p23 localized to the perinuclear Golgi region in all cell types, the distribution of the two proteins was clearly different even in the absence of nocodazole. It is not clear to what extent overlapping signal (yellow) is due to colocalization of the proteins or to superimposition of two close but separated fluorescent signals. Bar, 5 μm.
Mentions: Electron microscopy studies have localized COP I to the Golgi complex and to the transitional area between ER and Golgi (Oprins et al., 1993; Griffiths et al., 1995). Moreover, Sohn et al. (1996) have reported that rabbit p23 is enriched in COP I–coated vesicles and that p23 cytoplasmic tail peptides have the ability to recruit COP I, leading to the proposal that p23 represents a Golgi-specific receptor for COP I that is involved in the formation of COP I–coated vesicles. We therefore compared the localization of COP I with that of p23. Although both p23 and COP I exhibited a characteristic Golgi-like, perinuclear localization in different cell types, the distribution of the two proteins did not coincide (Fig. 8, CHO, Vero, and HeLa). This differential distribution was also evident after nocodazole-induced formation of dispersed Golgi stacks (Fig. 8, HeLa [+nocodazole]).

Bottom Line: Moreover, we find that p23 cytoplasmic domain is not involved in COP I membrane recruitment.Our data demonstrate that microinjected antibodies against the cytoplasmic tail of p23 inhibit G protein transport from the cis-Golgi network/ intermediate compartment to the cell surface, suggesting that p23 function is required for the transport of transmembrane cargo molecules.These observations together with the fact that p23 is a highly abundant component in the intermediate compartment, lead us to propose that p23 contributes to membrane structure, and that this contribution is necessary for efficient segregation and transport.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Geneva, 1211 Geneva 4, Switzerland.

ABSTRACT
Here, we report the localization and characterization of BHKp23, a member of the p24 family of transmembrane proteins, in mammalian cells. We find that p23 is a major component of tubulovesicular membranes at the cis side of the Golgi complex (estimated density: 12,500 copies/micron2 membrane surface area, or approximately 30% of the total protein). Our data indicate that BHKp23-containing membranes are part of the cis-Golgi network/intermediate compartment. Using the G protein of vesicular stomatitis virus as a transmembrane cargo molecule, we find that p23 membranes are an obligatory station in forward biosynthetic membrane transport, but that p23 itself is absent from transport vesicles that carry the G protein to and beyond the Golgi complex. Our data show that p23 is not present to any significant extent in coat protein (COP) I-coated vesicles generated in vitro and does not colocalize with COP I buds and vesicles. Moreover, we find that p23 cytoplasmic domain is not involved in COP I membrane recruitment. Our data demonstrate that microinjected antibodies against the cytoplasmic tail of p23 inhibit G protein transport from the cis-Golgi network/ intermediate compartment to the cell surface, suggesting that p23 function is required for the transport of transmembrane cargo molecules. These observations together with the fact that p23 is a highly abundant component in the intermediate compartment, lead us to propose that p23 contributes to membrane structure, and that this contribution is necessary for efficient segregation and transport.

Show MeSH
Related in: MedlinePlus