Fig. 6

pT172Ab recognized human pT181 GAP-43, which corresponds to rodent pT172. a pT172Ab (pT172) reacted with expressed human GAP-43 in transfected HEK 293 T cells, as well as GAP-43 expressed from mouse cDNA (Fig. 1a). This reactivity is probably due to phosphorylation of T181 in human GAP-43 (see Additional file 6: Fig. S5). HEK 293T cells were transiently transfected with human GAP-43-EGFP, GAP-43 T181A-EGFP-tagged vector, or N1-EGFP (control vector). In each lane, 28 μg of protein was loaded. b Human GAP-43 T181 was phosphorylated by JNK. The pT172Ab reaction was blocked by inhibitors of JNK (SP600125 (20 μM), inh. V (10 μM), or inh. XVI (1 μM)) for 3 h. Inhibitors of other MAPK signaling pathways (MEK1/2 inhibitor, but specific as an ERK inhibitor at this concentration: U0126 (5 μM)) and p38 inhibitor: SP203520 (5 μM)) did not affect this phosphorylation. Normalized intensities were measured using pT172Ab, and as a control, the band intensity of untreated cell was used (0.88% DMSO as a control, equal to the volume of DMSO in SP600125). n = 5; means ± SD. One-way ANOVA followed by a post hoc test using the Bonferroni method. **p < 0.01; ****p < 0.0001. c pT172Ab recognized the growth cones and the axons of differentiated neurons derived from hiPSCs, as well as those of mouse (see Fig. 1c). pT172Ab immunoreactivity was colocalized with that of GAP-43 Ab. Scale bar: 30 μm