The immunomodulatory beta-galactoside-specific lectin from mistletoe: partial sequence analysis, cell and tissue binding, and impact on intracellular biosignalling of monocytic leukemia cells.
Nanogram quantities of the beta-galactoside-specific lectin from mistletoe (ML-I) that is composed of two different types of subunits exhibit immunomodulatory potency and enhance cytokine secretion in vitro and in vivo. Partial sequence analysis of the carbohydrate-binding B chain revealed a ragged N-terminus and overall homologies to the B subunit of Ricin D and Ricin E. Two evolutionarily neutral substitutions were apparent in the otherwise identical N-terminal sequences of the two toxic chains within the lectin preparation. On the basis of the influence of chemical modification by group-specific reagents on ligand binding, the lectin was biotinylated with biotinyl-N-hydroxysuccinimide ester to allow monitoring of cell binding. Monocytic leukemia cells (THP-1) specifically bound the lectin with positive cooperativity at low lectin concentrations. Radiolabelled lectin could be found in several organs and in an experimental solid tumor in biodistribution in mice. Its presence in a notable amount in spleens is especially noteworthy with respect to the already reported immunomodulation. To determine intracellular responses that precede the lectin-dependent augmentation of cytokine secretion, phosphorylation of proteins and phospholipids as well as Ca(2+)-mobilization were assessed in THP-1 cells. Quantitative increases of [32P]-phosphate incorporation were determined for a 28 kDa protein and for phosphatidylinositol-4,5-biphosphate. Similarly, the fluorescence activity of the intracellular Ca(2+)-indicator fluo-3 is elevated by approximately 25% after lectin treatment. Apparently, cell binding of the lectin is followed by modulation of biosignalling processes.
Gabius
H J
HJ
Abteilung Glykobiochemie und Angewandte Tumorlektinologie, Institut für Pharmazeutische Chemie der Universität, Marburg, F.R.G.
Walzel
H
H
Joshi
S S
SS
Kruip
J
J
Kojima
S
S
Gerke
V
V
Kratzin
H
H
Gabius
S
S
D000595:Amino Acid Sequence; D000818:Animals; D005075:Biological Evolution; D001710:Biotin; D001921:Brain / Q000378:metabolism; D002286:Carcinoma, Ehrlich Tumor / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D005260:Female; D005697:Galactosides / Q000032:analysis; D006801:Humans; D046911:Macromolecular Substances; D008297:Male; D051379:Mice; D014784:Mistletoe; D008969:Molecular Sequence Data; D010920:Placenta / Q000378:metabolism; D028321:Plant Preparations; D010940:Plant Proteins; D010946:Plants, Medicinal; D011247:Pregnancy; D054790:Ribosome Inactivating Proteins, Type 2; D012689:Sequence Homology, Nucleic Acid; D014018:Tissue Distribution; D014118:Toxins, Biological / Q000737:chemistry* / Q000493:pharmacokinetics / Q000494:pharmacology
D005697:Galactosides; D046911:Macromolecular Substances; D028321:Plant Preparations; D010940:Plant Proteins; D054790:Ribosome Inactivating Proteins, Type 2; D014118:Toxins, Biological; C007816:beta-galactoside; C488862:mistletoe lectin I; C009674:ribosome inactivating protein, Viscum; D001710:Biotin