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A pore-forming toxin interacts with a GPI-anchored protein and causes vacuolation of the endoplasmic reticulum.

Abrami L, Fivaz M, Glauser PE, Parton RG, van der Goot FG - J. Cell Biol. (1998)

Bottom Line: Our data indicate that the protoxin binds to an 80-kD glycosyl-phosphatidylinositol (GPI)-anchored protein on BHK cells, and that the bound toxin is associated with specialized plasma membrane domains, described as detergent-insoluble microdomains, or cholesterol-glycolipid "rafts." We show that the protoxin is then processed to its mature form by host cell proteases.Strikingly, we found that the toxin causes dramatic vacuolation of the ER, but does not affect other intracellular compartments.Our data indicate that binding of proaerolysin to GPI-anchored proteins and processing of the toxin lead to oligomerization and channel formation in the plasma membrane, which in turn causes selective disorganization of early biosynthetic membrane dynamics.

View Article: PubMed Central - PubMed

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

ABSTRACT
In this paper, we have investigated the effects of the pore-forming toxin aerolysin, produced by Aeromonas hydrophila, on mammalian cells. Our data indicate that the protoxin binds to an 80-kD glycosyl-phosphatidylinositol (GPI)-anchored protein on BHK cells, and that the bound toxin is associated with specialized plasma membrane domains, described as detergent-insoluble microdomains, or cholesterol-glycolipid "rafts." We show that the protoxin is then processed to its mature form by host cell proteases. We propose that the preferential association of the toxin with rafts, through binding to GPI-anchored proteins, is likely to increase the local toxin concentration and thereby promote oligomerization, a step that it is a prerequisite for channel formation. We show that channel formation does not lead to disruption of the plasma membrane but to the selective permeabilization to small ions such as potassium, which causes plasma membrane depolarization. Next we studied the consequences of channel formation on the organization and dynamics of intracellular membranes. Strikingly, we found that the toxin causes dramatic vacuolation of the ER, but does not affect other intracellular compartments. Concomitantly we find that the COPI coat is released from biosynthetic membranes and that biosynthetic transport of newly synthesized transmembrane G protein of vesicular stomatitis virus is inhibited. Our data indicate that binding of proaerolysin to GPI-anchored proteins and processing of the toxin lead to oligomerization and channel formation in the plasma membrane, which in turn causes selective disorganization of early biosynthetic membrane dynamics.

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The distributions of the transferrin receptor (Trf-R),  the KDEL receptor ERD2 (ERD2), mannosidase II (Man II),  and the mannose-6-phosphate receptor (MPR) are not affected  by proaerolysin in BHK cells. Cells were incubated in the presence or absence of 0.38 nM proaerolysin either for 50 min (MPR)  or 1 h (Trf-R, ERD2, and Man II) at 37°C, and then processed for  immunofluorescence as described in Materials and Methods. Bar:  (Trf-R) 20 μm; (ERD2, Man II, and MPR) 13 μm.
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Figure 10: The distributions of the transferrin receptor (Trf-R), the KDEL receptor ERD2 (ERD2), mannosidase II (Man II), and the mannose-6-phosphate receptor (MPR) are not affected by proaerolysin in BHK cells. Cells were incubated in the presence or absence of 0.38 nM proaerolysin either for 50 min (MPR) or 1 h (Trf-R, ERD2, and Man II) at 37°C, and then processed for immunofluorescence as described in Materials and Methods. Bar: (Trf-R) 20 μm; (ERD2, Man II, and MPR) 13 μm.

Mentions: To identify the origin of the proaerolysin-induced vacuoles, we investigated whether they contained the transferrin receptor, a well-established marker of early endosomes (Hopkins and Trowbridge, 1983; Trowbridge et al., 1993) or rab7, a small GTPase found on late endosomal compartments (Chavrier et al., 1990; Feng et al., 1995; Méresse et al., 1995). The transferrin receptor (see Fig. 10) and rab7 (not shown) were clearly absent from the vacuoles, indicating that these did not originate from compartments of the endocytic pathway.


A pore-forming toxin interacts with a GPI-anchored protein and causes vacuolation of the endoplasmic reticulum.

Abrami L, Fivaz M, Glauser PE, Parton RG, van der Goot FG - J. Cell Biol. (1998)

The distributions of the transferrin receptor (Trf-R),  the KDEL receptor ERD2 (ERD2), mannosidase II (Man II),  and the mannose-6-phosphate receptor (MPR) are not affected  by proaerolysin in BHK cells. Cells were incubated in the presence or absence of 0.38 nM proaerolysin either for 50 min (MPR)  or 1 h (Trf-R, ERD2, and Man II) at 37°C, and then processed for  immunofluorescence as described in Materials and Methods. Bar:  (Trf-R) 20 μm; (ERD2, Man II, and MPR) 13 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2140172&req=5

Figure 10: The distributions of the transferrin receptor (Trf-R), the KDEL receptor ERD2 (ERD2), mannosidase II (Man II), and the mannose-6-phosphate receptor (MPR) are not affected by proaerolysin in BHK cells. Cells were incubated in the presence or absence of 0.38 nM proaerolysin either for 50 min (MPR) or 1 h (Trf-R, ERD2, and Man II) at 37°C, and then processed for immunofluorescence as described in Materials and Methods. Bar: (Trf-R) 20 μm; (ERD2, Man II, and MPR) 13 μm.
Mentions: To identify the origin of the proaerolysin-induced vacuoles, we investigated whether they contained the transferrin receptor, a well-established marker of early endosomes (Hopkins and Trowbridge, 1983; Trowbridge et al., 1993) or rab7, a small GTPase found on late endosomal compartments (Chavrier et al., 1990; Feng et al., 1995; Méresse et al., 1995). The transferrin receptor (see Fig. 10) and rab7 (not shown) were clearly absent from the vacuoles, indicating that these did not originate from compartments of the endocytic pathway.

Bottom Line: Our data indicate that the protoxin binds to an 80-kD glycosyl-phosphatidylinositol (GPI)-anchored protein on BHK cells, and that the bound toxin is associated with specialized plasma membrane domains, described as detergent-insoluble microdomains, or cholesterol-glycolipid "rafts." We show that the protoxin is then processed to its mature form by host cell proteases.Strikingly, we found that the toxin causes dramatic vacuolation of the ER, but does not affect other intracellular compartments.Our data indicate that binding of proaerolysin to GPI-anchored proteins and processing of the toxin lead to oligomerization and channel formation in the plasma membrane, which in turn causes selective disorganization of early biosynthetic membrane dynamics.

View Article: PubMed Central - PubMed

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

ABSTRACT
In this paper, we have investigated the effects of the pore-forming toxin aerolysin, produced by Aeromonas hydrophila, on mammalian cells. Our data indicate that the protoxin binds to an 80-kD glycosyl-phosphatidylinositol (GPI)-anchored protein on BHK cells, and that the bound toxin is associated with specialized plasma membrane domains, described as detergent-insoluble microdomains, or cholesterol-glycolipid "rafts." We show that the protoxin is then processed to its mature form by host cell proteases. We propose that the preferential association of the toxin with rafts, through binding to GPI-anchored proteins, is likely to increase the local toxin concentration and thereby promote oligomerization, a step that it is a prerequisite for channel formation. We show that channel formation does not lead to disruption of the plasma membrane but to the selective permeabilization to small ions such as potassium, which causes plasma membrane depolarization. Next we studied the consequences of channel formation on the organization and dynamics of intracellular membranes. Strikingly, we found that the toxin causes dramatic vacuolation of the ER, but does not affect other intracellular compartments. Concomitantly we find that the COPI coat is released from biosynthetic membranes and that biosynthetic transport of newly synthesized transmembrane G protein of vesicular stomatitis virus is inhibited. Our data indicate that binding of proaerolysin to GPI-anchored proteins and processing of the toxin lead to oligomerization and channel formation in the plasma membrane, which in turn causes selective disorganization of early biosynthetic membrane dynamics.

Show MeSH
Related in: MedlinePlus