Molecular cloning and expression of cDNA encoding human macrophage C-type lectin. Its unique carbohydrate binding specificity for Tn antigen.
A human macrophage calcium-dependent (C-type) lectin cDNA clone was obtained from a library derived from IL-2-treated peripheral blood monocytes. The cDNA cloning was based on the structural homology to hepatic asialoglycoprotein receptors. The nucleotide sequence of this cDNA clone was homologous to those of the galactose- and N-acetylgalactosamine-specific C-type macrophage lectins of rodents. In the putative carbohydrate recognition domain, deduced amino acid sequence revealed 60 and 63% homology to galactose- and N-acetylgalactosamine-specific C-type macrophage lectins of mice and rats, respectively. The cDNA clone was ligated into a mammalian expression vector and transfected into COS-1 cells. In the lysates of these cells, an MR 38,000 component, which bound to galactose-Sepharose, was identified after electrophoretic separation by its interaction with polyclonal antisera against synthetic polypeptides representing a portion of the carbohydrate recognition domain. The carbohydrate-binding specificity of the recombinant macrophage lectin was investigated by comparing elution profiles of various glycopeptides having defined carbohydrate structures from immobilized macrophage lectins. When N-terminal octapeptides from human glycophorin A that bore NeuAc alpha 2-3 Gal beta 1-3 (NeuAc alpha 2-6) GalNAc and its sequentially deglycosylated derivatives were compared, glycopeptides carrying three constitutive GalNAc-Ser/Thr (Tn Ag) strongly bound to the recombinant human macrophage lectin. This is the first study to demonstrate that human macrophage cell surface lectin recognizes Tn Ag, a well-known human carcinoma-associated epitope.
Suzuki
N
N
Department of Cancer Biology and Molecular Immunology, Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
Yamamoto
K
K
Toyoshima
S
S
Osawa
T
T
Irimura
T
T
D000595:Amino Acid Sequence; D015295:Antigens, Tumor-Associated, Carbohydrate / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D001212:Asialoglycoproteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D002118:Calcium / Q000502:physiology*; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D000939:Epitopes / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D037102:Lectins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D037181:Lectins, C-Type; D008264:Macrophages / Q000737:chemistry / Q000276:immunology*; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry
D015295:Antigens, Tumor-Associated, Carbohydrate; D001212:Asialoglycoproteins; D002352:Carrier Proteins; D000939:Epitopes; D037102:Lectins; D037181:Lectins, C-Type; D008565:Membrane Proteins; D011994:Recombinant Proteins; C040286:Tn antigen; D002118:Calcium