Cdc42 (A,B) and Rac1 (C, D) are activated at lower levels in cells transfected with agrin GAG chain mutants than with wild-type agrin. Cells were co-transfected with wild type Cdc42 or Rac1 and GFP, or TM-agrin-GFP, SI, SII or SI+II. After 24 h, cells were lysed and the activation of Rac1 and Cdc42 were measured by a pull-down assay. Total over-expressed Cdc42 or Rac1 protein was visualized by Western blotting using an AU5 antibody. Graphs show mean activation level and standard error (n = 3 experiments).