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Purification and characterization of the apical plasma membrane of the rat pancreatic acinar cell

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Summary

A method is described for the rapid purification of the apical plasma membrane from the rat pancreatic acinar cell. It makes use of wheat germ agglutinin affinity chromatography to selectively bind vesicles with N-acetyl glucosamine present at their surface. Particular conditions (150 mm NaCl) had then to be used to keep membrane vesicles in the coveted orientation, i.e. as right-side-out vesicles. Due to its specific apical location in many epithelial cells, γ-glutamyltranspeptidase was chosen to monitor the purification procedure. The final fraction was enriched in γ-glutamyltranspeptidase by a factor of 75 relative to the homogenate. Na,K-ATPase, a strict basolateral membrane marker, was not detectable in the fraction. No membranes originating from other compartments, more particularly expected from zymogen granules, or from other cell types, did contaminate the preparation. As expected for an epithelial cell apical plasmalemma, lipid composition showed a very high ratio of glycolipids (37.5%). The absence of membrane-bound GP-2, and the exceptionally high specific activity of γ-glutamyltranspeptidase suggest that the apical membrane would not be made up by the exocytosis of secretory granule, but instead by the fusion of specialized secretory vesicles very likely originating from the constitutive secretory pathway. In conclusion, this report describes a method of obtaining a fraction highly enriched in the secretory apex of the pancreatic exocrine cell that would be directly involved in exocytosis with zymogen granules and also in local anion transport.

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Authors and Affiliations

  1. Centre de Recherche sur les Mécanismes de Sécrétion, Département de Biologie, Faculté des Sciences, Université de Sherbrooke, J1K 2R1, Sherbrooke, Québec, Canada

    Emmanuel Paul, Yves Hurtubise & Denis LeBel

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  1. Emmanuel Paul
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  2. Yves Hurtubise
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  3. Denis LeBel
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Additional information

The authors would like to thank Dr. Andrew W. Shyjan (Yale University, New Haven, CT) for his kind gift of anti-Na,K-ATPase β1 subunit, Dr. Yannick Laperche (INSERM, Hôpital Mondor, Paris) for his gift of anti-γ-GT, and finally Mr. Gilles P. Grondin for the production of antibodies against amylase. D.L. is supported by NSERC of Canada, FCAR of Québec, and the Canadian Cystic Fibrosis Foundation.

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Paul, E., Hurtubise, Y. & LeBel, D. Purification and characterization of the apical plasma membrane of the rat pancreatic acinar cell. J. Membarin Biol. 127, 129–137 (1992). https://doi.org/10.1007/BF00233285

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  • DOI: https://doi.org/10.1007/BF00233285

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