Figure 2. p45 interferes with the p75-NgR interaction and signaling.

(A) Lysates from HEK293 cells co-transfected with vectors expressing p45, p75, and Flag-tagged human NogoR (Flag-hNgR) were immunoprecipitated with anti-p75 antibodies and probed with anti-Flag antibodies. The results showed that p75 and hNgR form a complex and presence of p45 reduces the formation of p75/hNgR complex. (B) Co-transfection with varying amount of p45-expressing vectors markedly reduced p75/hNgR complex formation in a concentration-dependent fashion. (C) Quantification of the increase of RhoA activity after MAG-Fc treatment. In wild-type (WT) CGN cultures, when treated with MAG-Fc, the RhoA activity increased by 50%. However, in CGN cultures derived from Thy1-p45 transgenic mice, the increase in RhoA activity induced by MAG-Fc treatment is completely abolished. The value is expressed as percent over control. The value is derived from three independent experiments. * p<0.05, Student's t test. The data can be found in Table S1. (D) CGNs were seeded on glass coverslips coated with inhibitory substrates, grown for 14–18 h, and immunofluorescently stained with Tuj1 (Red) and anti-p45 (Green) antibodies. Scale bar, 50 µm. (E) Quantitative analysis of neurite length from the outgrowth assay, using Nogo66-GST, myelin, or HEK293 cells expressing MAG as inhibitory substrates. The data are represented as mean ± SEM (* p<0.001) and can be found in Table S1. Overexpression of p45 promoted neurite outgrowth of CGNs on inhibitory substrates.