Immunocytochemical localization of alpha2,3(N)-sialyltransferase (ST3Gal III) in cell lines and rat kidney tissue sections: evidence for golgi and post-golgi localization |
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Authors: | Burger, PC Lotscher, M Streiff, M Kleene, R Kaissling, B Berger, EG |
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Affiliation: | Institute of Physiology, University of Zurich, Switzerland. |
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Abstract: | Sialylation is a biosynthetic process occurring in the trans compartmentsof the Golgi apparatus. Corresponding evidence is based on localization andbiochemical studies of alpha2, 6(N)-sialyltransferase (ST6Gal I) aspreviously reported. Here we describe generation and characterization ofpolyclonal antibodies to recombinant rat alpha2,3(N)-sialyltransferase(ST3Gal III) expressed as a soluble enzyme in Sf9 cells or as abeta-galactosidase-human-ST3Gal III fusion- protein from E.coli ,respectively. These antibodies were used to localize ST3Gal III byimmunofluorescence in various cell lines and rat kidney tissue sections. Intransiently transfected COS cells the antibodies directed to solublesialyltransferase or the sialyltransferase portion of the fusion-proteinonly recognized the recombinant antigen retained in the endoplasmicreticulum. However, an antibody fraction crossreactive withbeta-galactosidase recognized natively expressed ST3Gal III which was foundto be colocalized with beta1, 4-galactosyltransferase in the Golgiapparatus of several cultured cell lines. Antibodies affinity purified onthe beta- galactosidase-ST3Gal III fusion-protein column derived from bothantisera have then been used to localize the enzyme in perfusion-fixed ratkidney sections. We found strong staining of the Golgi apparatus of tubularepithelia and a brush-border-associated staining which colocalized withcytochemical staining of the H+ATPase. This subcellular localization wasnot observed for ST6Gal I which localized to the Golgi apparatus. Thesedata show colocalization in the Golgi apparatus and different post-Golgidistributions of the two sialyltransferases. |
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