FIGURE 7.

Loss of ERM-mediated PKA anchoring converts netrin-induced growth cone attraction to repulsion. A, IMR-32 cells serially transfected with shERM plasmids and rescue plasmids expressing either wild-type (WT) or AKAP-deficient (L421P) radixin were plated onto poly-l-lysine-coated coverslips in serum-free medium containing 5 μm all-trans retinoic acid for 12 h. Phase contrast images were acquired before and 30 min after exposure to a gradient of either BSA (Control) or netrin-1 (5 μg/ml in the pipette; position indicated by the arrowheads) and processed using a custom ImageJ macro to generate protrusion-retraction analysis maps (PRAM). B and C, directional growth cone responses were quantified by calculating the protrusion or retraction index toward (Proximal) or away from (Distal) to the gradient source. P, protrusion; R, retraction; O, overlap. Summed data are plotted in C as the mean ± S.D. for at least five cells from two separate experiments.