Mutation of GrB cleavage site inhibited nuclear translocation of FGFR1. (A) Site-directed mutagenesis on wild-type (WT) FGFR1 changed the guanine nucleotide at position 1963 to adenine (g1963a; using sequence available from GenBank/EMBL/DDBJ under accession no. NM_023106.2) and thereby amino acid 432 from aspartic acid to alanine (D432N), abolishing the GrB cleavage site in Mutant FGFR1. (B) Confocal analysis of MCF-7 cells transiently transfected (48 h) with WT (VSAD) or mutant (VSAN) FGFR1b revealed that mutant (VSAN) FGFR1b did not translocate to the nucleus. Bar, 50 µm. (C) Transwell migration assays showed that, under control conditions, MCF-7 cells transfected with WT FGFR1b (VSAD) migrated more compared with empty vector (pcDNA4/TO)– and mutant FGFR1b (VSAN)–transfected cells. FGF-10 stimulation increased migration in cells transfected with WT FGFR1b (VSAD) but did not affect MCF-7 cells transfected with mutant FGFR1b (VSAN). *, P ≤ 0.05 (paired Student’s t test and ANOVA). Error bars show means ± SEM.