FIGURE 6.

Neurite outgrowth induced by NGF is blocked by overexpression of dominant-negative ERK5 and MEK5A mutants. A, PC12 cells were co-transfected with GFP and empty vector or ERK5KD. The cells were incubated with NGF (100 ng/ml) or Bt₂cAMP (dbcAMP) (0.5 mm) for a day. The nuclei were stained with Hoechst-33258 (blue), and morphological changes of GFP-positive cells were observed under a fluorescence microscope. Scale bar, 100 μm. Neurite length of GFP-positive PC12 cells was measured as described under “Experimental Procedures.” Values represent the means ± S.E. (n = 3). NGF and Bt₂cAMP significantly increased neurite length (*, p < 0.05), and the effect of NGF was reversed by ERK5KD (#, p < 0.05). B, parental PC12 cells or PC12 cells stably overexpressing MEK5A mutant were incubated with NGF (100 ng/ml) or Bt₂cAMP (0.5 mm) for a day. The nuclei were stained with Hoechst-33258, and the morphological changes were observed. Scale bar, 100 μm. Neurite length of PC12 cells was measured as described under “Experimental Procedures.” Values represent the means ± S.E. (n = 3). NGF and Bt₂cAMP significantly increased neurite length (*, p < 0.05), and the effect of NGF was reversed by MEK5A overexpression (#, p < 0.05).