ACTIVATED RAT MACROPHAGES PRODUCE A GALECTIN-1-LIKE PROTEIN THAT INDUCES APOPTOSIS OF T-CELLS - BIOCHEMICAL AND FUNCTIONAL-CHARACTERIZATION

Galectins, a family of closely related beta-galactoside-binding protei ns, show specific immunomodulatory properties. We have recently identi fied the presence of a galectin-like protein in rat peritoneal macroph ages by means of a cross-reactivity with a polyclonal Ab raised agains t a galectin purified from adult chicken liver. Galectin expression wa s up-regulated in inflammatory and activated macrophages, revealing a significant increase in phorbol ester-and formylmethionine oligopeptid e-treated cells. In an attempt to further explore its functional signi ficance, rat macrophage galectin was purified from activated macrophag es by a single-step affinity chromatography on a lactosyl-Sepharose ma trix. The eluted fraction was resolved as a single protein band of sim ilar to 15,000 Da by SDS-PAGE that immunoreacted strongly with the ant i-chicken galectin serum. Gel filtration studies revealed that the pro tein behaved like a dimer under native conditions, and saccharides bea ring a beta-D-galactoside configuration were able to inhibit the hemag glutinating activity displayed by the purified galectin, In agreement with its isoelectric point of similar to 4.8, the amino acid analysis showed a definitive acidic pattern. Internal amino acid sequencing of selected peptides obtained by proteolytic cleavage revealed that this carbohydrate-binding protein shares ail the absolutely preserved and c ritical residues found in other members of the mammalian galectin-1 su bfamily, Finally, biochemical and ultrastructural evidence, obtained b y genomic DNA fragmentation and transmission electron microscopy, are also provided to show its potential implications in the apoptotic prog ram of T cells, This effect was quantified by using the terminal deoxy nucleotidyl transferase-mediated dUTP biotin nick end-labeling assay a nd was found to be associated to the specific carbohydrate-binding pro perties of galectin.