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

Galectin-3 is a member of the beta-galactoside-binding protein family shown to be involved in tumor progression and metastasis, It has a unique primar y structure consisting of three domains: a 12-amino acid leader sequence co ntaining a casein kinase I serine phosphorylation site, which is preceded b y a collagenase-sensitive Pro-Gly-rich motif, and a COOH-terminal half enco mpassing the carbohydrate-binding site. To study the functional role of the unusual leader sequence of galectin-3, a mutant cDNA that causes an 11-ami no 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 te rminus resulted in abolition of the secretion of truncated galectin-3, Loss of nuclear localization, and reduced carbohydrate-mediated functions compa red with the wild-type protein. When green fluorescent protein was fused to the galectin-3 leader sequence and transiently transfected into BT-549 cel ls, the uniform cellular distribution of native green fluorescent protein w as 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 Ser(6), two point muta tions were created at this serine: Ser(6)-->Ala and Ser(6)-->Glu. No obviou s difference was observed in cellular localization between wild-type and Se r(6)-mutated transfectants. These results suggest a structural role for the NH2 terminus leader motif of galectin-3 in determining its cellular target ing and biological functions independent of phosphorylation.