Localization of galactosyl- and sialyltransferase by immunofluorescence: evidence for different sites.

Abstract

Polyclonal rabbit antisera against soluble human milk galactosyltransferase and bovine colostrum sialyltransferase were used to localize by indirect immunofluorescence the respective intracellular enzymes in primary cultures from bovine fetal kidneys and established cell lines of human and bovine fibroblasts. Staining for galactosyltransferase was juxtanuclear and crescent shaped in epitheloid cells; a similar staining, occasionally perinuclear and sparsely distributed in the cytoplasm, was found in fibroblasts. In contrast, staining for sialyltransferase in epitheloid kidney cells derived from the same primary culture was observed predominantly in cytoplasmic vesicles that were spread over the whole cytoplasm. Sialyltransferase-positive vesicles had a similar distribution in fibroblasts and often appeared concentrated around an unstained Golgi area. Thus, in both cell types galactosyl-and sialyltransferase were localized in different subcellular compartments. Since both galactosyl-and sialyltransferase participate in formation ofthe terminal glycan NeuAc(a2--6)Gal(fi1-+4)GlcNAc (Neu, neuraminic acid) present in many N-glycosidic complex types of glycans, different subcellular compartments for these enzymes support a model of functional compartmentalization of the Golgi apparatus that is compatible with an assembly-line model for glycan chain elongation and termination.

A role for GRIP domain proteins and/or their ligands in structure and function of the trans Golgi network

“…Stable transfectants of the CHO cell variant TRVb-1 expressing Tac chimeric proteins TGG and TTF (Ghosh et al, 1998;Mallet and Maxfield, 1999) (Prescott et al, 1997); sheep anti-TGN46, Serotec, Oxford, UK)]; cation-independent mannose 6-phosphate receptor (mAb αMPR300, Affinity BioReagents, Golden, CO); Tac (Santini et al, 1998); golgin-97 (rb αgolgin-97 and mAb αgolgin-97, E. K. L. Chan, Scripps Res. Inst., La Jolla, CA); giantin (Linstedt and Hauri, 1993); Rab6 (C-19 rb αRab6, Santa Cruz Biotechnology, Santa Cruz, CA); ERGIC-53 (Schweizer et al, 1988); mannosidase II (Moremen and Touster, 1985); galactosyltransferase (Berger and Hesford, 1985); MG160 (Gonatas et al, 1989); AP-1 (Ahle et al, 1988); Lamp1 (Carlsson et al, 1988;Mane et al, 1989); Lamp2 (Mane et al, 1989); CD63 (mAb αCD63, Beckman Coulter, Fullerton, CA); transferrin receptor (mAb B3/25, Roche Molecular Biochemicals); T7 epitope (mAb from Novagen, Madison, WI). mAbs to EEA1, human tGolgin-1 (p230), GM130, p115 and syntaxin 6 were from Becton Dickenson/ Transduction Laboratories (San Diego, CA).…”

tGolgin-1 (p230, golgin-245) modulates Shiga-toxin transport to the Golgi and Golgi motility towards the microtubule-organizing centre

“…Only the portion of the gel corresponding to full-length tGolgin-1 (260 kDa) is shown; the rest of the lanes were blank. tGolgin-1 regulates Golgi link to motors anti-golgin-97 and monoclonal antibody (mAb) anti-golgin-97, E. K. L. Chan (Scripps Research Institute, La Jolla, CA) (Griffith et al, 1997); mAb anti-giantin, H.-P. Hauri (University of Basel, Switzerland) (Linstedt and Hauri, 1993); rabbit anti-β-1,4-galactosyl transferase (GalT), E. Berger (University of Zurich, Switzerland) (Berger and Hesford, 1985); mAb anti-Lamp1 (H4A3), Developmental Studies Hybridoma Bank, Iowa City, IA; anti-CD63 (mAb against CD63), Beckman Coulter, Fullerton, CA; anti-tubulin (mAb YL1/2), Accurate Chemical, Westbury, NY; anti-transferrin receptor (mAb B3/25), Roche Molecular Biochemicals, Indianapolis, IN. mAbs against human tGolgin-1 (p230), EEA1, GM130, p115 and GS28 were from Becton Dickinson/Pharmingen (San Diego, CA).…”

Muscle cell components dictate hepatocyte gene expression and the distribution of the Golgi apparatus in heterokaryons.

“…Individual cells were analyzed to determine whether hepatocyte nuclei contained in a heterokaryon retained a Golgi distribution that was typical of hepatocytes or changed to one typical of striated muscle. For this purpose we used an antibody to a Golgi enzyme, galactosyl transferase, which is localized in the trans cistemae of the Golgi stack and readily visualized by immunofluorescence (Berger et al 1981;Berger and Hesford 1985;Lucocq et al 1987).…”