The NH₂ terminus of galectin-3 governs cellular compartmentalization and functions in cancer cells

Galectin-3 is a member of the β-galactoside-binding protein family shown to be involved in tumor progression and metastasis. It has a unique primary structure consisting of three domains: a 12-amino acid leader sequence containing a casein kinase I serine phosphorylation site, which is preceded by a collagenase-sensitive Pro-Gly-rich motif, and a COOH-terminal half encompassing the carbohydrate-binding site. To study the functional role of the unusual leader sequence of galectin-3, a mutant cDNA that causes an 11-amino acid deletion in the NH₂-terminal region was generated and expressed in galectin-3-null BT-549 human breast carcinoma cells. Deletion of the NH₂ terminus resulted in abolition of the secretion of truncated galectin-3, loss of nuclear localization, and reduced carbohydrate-mediated functions compared with the wild-type protein. When green fluorescent protein was fused to the galectin-3 leader sequence and transiently transfected into BT-549 cells, the uniform cellular distribution of native green fluorescent protein was changed mainly to a nuclear pattern. To further investigate whether the functional changes observed in a galectin-3 with the 11 NH₂- terminal amino acids deleted were due to loss of phosphorylation at Ser⁶, two point mutations were created at this serine: Ser⁶→Ala and Ser⁶→Glu. No obvious difference was observed in cellular localization between wild-type and Ser⁶-mutated transfectants. These results suggest a structural role for the NH₂ terminus leader motif of galectin-3 in determining its cellular targeting and biological functions independent of phosphorylation.