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Galectin-4 and sulfatides in apical membrane trafficking in enterocyte-like cells.

Delacour D, Gouyer V, Zanetta JP, Drobecq H, Leteurtre E, Grard G, Moreau-Hannedouche O, Maes E, Pons A, André S, Le Bivic A, Gabius HJ, Manninen A, Simons K, Huet G - J. Cell Biol. (2005)

Bottom Line: Moreover, galectin-4 depletion altered the DRM association characteristics of apical proteins.Sulfatides with long chain-hydroxylated fatty acids, which were also enriched in DRMs, were identified as high-affinity ligands for galectin-4.Together, our data propose that interaction between galectin-4 and sulfatides plays a functional role in the clustering of lipid rafts for apical delivery.

View Article: PubMed Central - PubMed

Affiliation: Unité INSERM 560, 59045 Lille Cedex, France.

ABSTRACT
We have previously reported that 1-benzyl-2-acetamido-2-deoxy-alpha-D-galactopyranoside (GalNAc alpha-O-bn), an inhibitor of glycosylation, perturbed apical biosynthetic trafficking in polarized HT-29 cells suggesting an involvement of a lectin-based mechanism. Here, we have identified galectin-4 as one of the major components of detergent-resistant membranes (DRMs) isolated from HT-29 5M12 cells. Galectin-4 was also found in post-Golgi carrier vesicles. The functional role of galectin-4 in polarized trafficking in HT-29 5M12 cells was studied by using a retrovirus-mediated RNA interference. In galectin-4-depleted HT-29 5M12 cells apical membrane markers accumulated intracellularly. In contrast, basolateral membrane markers were not affected. Moreover, galectin-4 depletion altered the DRM association characteristics of apical proteins. Sulfatides with long chain-hydroxylated fatty acids, which were also enriched in DRMs, were identified as high-affinity ligands for galectin-4. Together, our data propose that interaction between galectin-4 and sulfatides plays a functional role in the clustering of lipid rafts for apical delivery.

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GalNAcα-O-bn decreases the apical localization of galectin-4. (A) Western blot of culture media, cytosol and membrane fractions of control and GalNAcα-O-bn–treated (14 d) cells after permeabilization with saponin, using anti–galectin-4 antibody. (B) Confocal microscopy with an anti–galectin-4 antibody on permeabilized or unpermeabilized control and GalNAcα-O-bn–treated (14 d) cells. Apical and xz sections are shown. Bars, 22 μm.
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fig2: GalNAcα-O-bn decreases the apical localization of galectin-4. (A) Western blot of culture media, cytosol and membrane fractions of control and GalNAcα-O-bn–treated (14 d) cells after permeabilization with saponin, using anti–galectin-4 antibody. (B) Confocal microscopy with an anti–galectin-4 antibody on permeabilized or unpermeabilized control and GalNAcα-O-bn–treated (14 d) cells. Apical and xz sections are shown. Bars, 22 μm.

Mentions: The distribution of galectin-4 in control and in GalNAcα-O-bn–treated cells was studied by analyzing the culture medium, the cytosolic and the total membrane fractions from saponin-permeabilized cells for the presence of galectin-4 (Fig. 2 A). In control cells, galectin-4 was found in the membrane fraction, whereas in GalNAcα-O-bn–treated cells, galectin-4 was found in the soluble cytosolic fraction. Neither the control cells nor the GalNAcα-O-bn–treated cells secreted detectable levels of galectin-4 to the culture medium.


Galectin-4 and sulfatides in apical membrane trafficking in enterocyte-like cells.

Delacour D, Gouyer V, Zanetta JP, Drobecq H, Leteurtre E, Grard G, Moreau-Hannedouche O, Maes E, Pons A, André S, Le Bivic A, Gabius HJ, Manninen A, Simons K, Huet G - J. Cell Biol. (2005)

GalNAcα-O-bn decreases the apical localization of galectin-4. (A) Western blot of culture media, cytosol and membrane fractions of control and GalNAcα-O-bn–treated (14 d) cells after permeabilization with saponin, using anti–galectin-4 antibody. (B) Confocal microscopy with an anti–galectin-4 antibody on permeabilized or unpermeabilized control and GalNAcα-O-bn–treated (14 d) cells. Apical and xz sections are shown. Bars, 22 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171948&req=5

fig2: GalNAcα-O-bn decreases the apical localization of galectin-4. (A) Western blot of culture media, cytosol and membrane fractions of control and GalNAcα-O-bn–treated (14 d) cells after permeabilization with saponin, using anti–galectin-4 antibody. (B) Confocal microscopy with an anti–galectin-4 antibody on permeabilized or unpermeabilized control and GalNAcα-O-bn–treated (14 d) cells. Apical and xz sections are shown. Bars, 22 μm.
Mentions: The distribution of galectin-4 in control and in GalNAcα-O-bn–treated cells was studied by analyzing the culture medium, the cytosolic and the total membrane fractions from saponin-permeabilized cells for the presence of galectin-4 (Fig. 2 A). In control cells, galectin-4 was found in the membrane fraction, whereas in GalNAcα-O-bn–treated cells, galectin-4 was found in the soluble cytosolic fraction. Neither the control cells nor the GalNAcα-O-bn–treated cells secreted detectable levels of galectin-4 to the culture medium.

Bottom Line: Moreover, galectin-4 depletion altered the DRM association characteristics of apical proteins.Sulfatides with long chain-hydroxylated fatty acids, which were also enriched in DRMs, were identified as high-affinity ligands for galectin-4.Together, our data propose that interaction between galectin-4 and sulfatides plays a functional role in the clustering of lipid rafts for apical delivery.

View Article: PubMed Central - PubMed

Affiliation: Unité INSERM 560, 59045 Lille Cedex, France.

ABSTRACT
We have previously reported that 1-benzyl-2-acetamido-2-deoxy-alpha-D-galactopyranoside (GalNAc alpha-O-bn), an inhibitor of glycosylation, perturbed apical biosynthetic trafficking in polarized HT-29 cells suggesting an involvement of a lectin-based mechanism. Here, we have identified galectin-4 as one of the major components of detergent-resistant membranes (DRMs) isolated from HT-29 5M12 cells. Galectin-4 was also found in post-Golgi carrier vesicles. The functional role of galectin-4 in polarized trafficking in HT-29 5M12 cells was studied by using a retrovirus-mediated RNA interference. In galectin-4-depleted HT-29 5M12 cells apical membrane markers accumulated intracellularly. In contrast, basolateral membrane markers were not affected. Moreover, galectin-4 depletion altered the DRM association characteristics of apical proteins. Sulfatides with long chain-hydroxylated fatty acids, which were also enriched in DRMs, were identified as high-affinity ligands for galectin-4. Together, our data propose that interaction between galectin-4 and sulfatides plays a functional role in the clustering of lipid rafts for apical delivery.

Show MeSH
Related in: MedlinePlus