The novel lectin KM+ detects a specific subset of mannosyl-glycoconjugates in the rat cerebellum.
KM+ is a D(+)mannose-specific lectin with a carbohydrate structure-affinity relationship different from those of most mannose-binding lectins. KM+ elicits carbohydrate-dependent biological effects in several mammalian cell types, but it has not yet been employed as a probe for the detection of its specific ligands. We show here for the first time the screening and partial identification of cerebellar mannosyl-glycoconjugates recognized by KM+, by means of lectin-histochemistry and lectin-blotting. Biotinylated KM+ stained most cellular structures in the adult rat cerebellum, particularly Purkinje cells bodies and the surface of granule cells, but not cellular processes. Capillaries in the choroid plexus were also strongly decorated, while blood vessels in the cerebellar parenchyma remained unstained. D(+)mannose, but not D(+)galactose, abolished the staining of all cerebellar structures. Higher inhibitory potencies were found for mannosyl-glycans such as mannotriose (man-alpha1,3-[man-alpha1,6]-man) and the biantennary heptasaccharide carried by the enzyme horseradish peroxidase. After separation of cerebellar proteins by SDS-PAGE, KM+ recognized three major unidentified mannosyl-glycoproteins of 132, 83 and 49 kDa. KM+ also detected high-Mw bands corresponding to the light and heavy chains of Type-I laminin, but not a 160-kDa cleavage product of laminin. We conclude that KM+ binds preferentially to a specific subset of mannose-containing glycoproteins in cerebellar tissue, thus being much more restricted than other mannose-specific lectins. KM+ can be used as a novel probe to screen the central nervous system for this specific subset of complex mannosyl-glycoconjugates.
Department of Pharmacology, School of Medicine, University of Campinas, Brazil. sateixeira@hcrp.fmrp.usp.br
Silvia A
SA
Teixeira
Mariano S
MS
Viapiano
Luciane
L
Ganiko
Maria Cristina
MC
Roque-Barreira
Antonio R
AR
Martins
D000818:Animals; D002531:Cerebellum / Q000737:chemistry*; D006023:Glycoproteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D007797:Laminin / Q000032:analysis; D008024:Ligands; D008358:Mannose / Q000032:analysis*; D037241:Mannose-Binding Lectins / Q000378:metabolism*; D051381:Rats
D006023:Glycoproteins; D007797:Laminin; D008024:Ligands; D037241:Mannose-Binding Lectins; C121346:laminin 1; D008358:Mannose