Isolation and immunohistochemical localization of a lectin-like molecule from the rat cerebellum.
A lectin with a mannose specificity was isolated from the cerebellum of young rats. The method of purification was based on the observation that during homogenization of the tissue, the lectin binds to a class of mannose-rich glycoproteins highly insoluble in Triton X-100. Sequential extractions in saline buffer devoid of, then containing, 0.5% Triton X-100 allowed the elimination of a great part of other proteins. Using the same buffer containing 0.5 M mannose, a specific class of protein can be solubilized. This fraction was enriched by affinity adsorption on insolubilized mannose-rich glycoproteins followed by specific detachment with mannose. One of the protein subunits, of molecular weight (MW) 130,000, was isolated by preparative gel electrophoresis. Upon re-electrophoresis, this compound gives two bands of MW 65,000 and 130,000, which appear to be a monomer and a dimer of a molecule called R1. Antibodies were raised against R1 which react with the monomer and the dimer and not against other proteins of the rat cerebellum. The immunohistochemical localization of this lectin was performed in cerebella of 20-day-old rats. The antigen is concentrated in endothelial cells and in large and intermediate size neurons (Purkinje, Golgi, basket and deep nuclei neurons). Granule cell bodies are lightly stained and no label at all was found in glial cells. At the level of electron microscopy, the antigen was found to be very concentrated in multivesicular bodies and lysosomes of large neurons, on parts of the endoplasmic reticulum, on some mitochondrial outer membranes and on the plasma membrane of the dendrites. The possible role of this lectin in cerebella of young rats is discussed in relation to its interaction with a specific class of mannose-rich glycoproteins.
D000818:Animals; D002531:Cerebellum / Q000032:analysis* / Q000378:metabolism; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000032:analysis; D006651:Histocytochemistry; D007120:Immunochemistry; D037102:Lectins / Q000276:immunology / Q000302:isolation & purification*; D008854:Microscopy, Electron; D051381:Rats; D011919:Rats, Inbred Strains
D006023:Glycoproteins; D037102:Lectins