FIGURE 8:

R1881 and NGF trigger association of a multiprotein complex and stimulate Rac-dependent neurite outgrowth. Quiescent PC12 cells were used. (A, B) Plastic-plated cells were left untreated or treated for 5 min with the R1881 (10 nM) or NGF (100 ng/ml) in the absence or presence of bicalutamide (Bic at 10 μM). Lysate proteins were immune-precipitated with C-19 anti-AR (A) or anti-Trk (B) antibodies or nonspecific immunoglobulin G (Ctrl Ab) as a control. Proteins in immune complexes were detected by Western blot, using the antibodies against the indicated proteins. The C-19 anti AR Ab was used to detect AR. (C) Cells on polylysine-coated coverslips were made quiescent and left untreated or treated for 24 h with R1881 (10 nM) or NGF (100 ng/ml) in the absence or presence of EHT (5 μM). Neurite outgrowth was analyzed by contrast phase microscopy and is expressed as percentage of total cells. Means and SEM are shown. Asterisks indicate significantly different values as compared with control (p < 0.05). (D) Plastic-plated cells were left untreated or treated for 5 min with 10 nM R1881 (top) or 100 ng/ml NGF (bottom) in the absence or presence of the indicated compounds. Bicalutamide (Bic) was used at 10 μM, S peptide at 1 nM, and IC87114 (IC) at 5 μM. Rac activation was analyzed by pull-down assay. Active (Rac-GTP) or total Rac (Rac) was detected by Western blot.