Exogenous gal-3 binding to CD45 reduces tyrosine phosphatase activity. (A) SUDHL-6 and SUDHL-9 cells were treated with 5μM gal-3 and intracellular tyrosine phosphatase activity determined spectrophotometrically. Exogenous gal-3 significantly decreased phosphatase activity in SUDHL-6 cells, but had no effect on phosphatase activity in SUDHL-9 cells (***P < .0001). (B) Gal-3 reduction of CD45 phosphatase activity is glycan dependent. Gal-3 was preincubated with 100mM lactose or sucrose or with buffer control for 15 minutes at 37°C before treatment of SUDHL-6 cells. Although treatment with gal-3 alone or gal-3 plus sucrose significantly reduced phosphatase activity (***P < .0001), no reduction in phosphatase activity was seen in cells treated with gal-3 plus lactose to block gal-3 binding. (C) Multimerization of gal-3 is required to reduce phosphatase activity. Gal-3C is the C-terminal domain of gal-3 that lacks the N-terminal peptide required for multimerization. SUDHL-6 cells were treated with gal-3 or gal-3C and assayed for phosphatase activity. Although gal-3 significantly reduced phosphatase activity (***P < .0001), gal-3C did not affect phosphatase activity. Values are mean ± SD for triplicates from 1 representative of 3 independent replicate experiments.