Figure 8.
NGF-activated CK2 participates in NGF/p75NTR signaling, leading to PTEN and GSK-3β phosphorylation and subsequent axonal growth. (A) CK2 activity was determined from cytosolic extracts obtained from cultured hippocampal neurons. Cultures were incubated for 30 min in the presence or absence of NGF, DMAT (a specific CK2 inhibitor), or NGF plus DMAT. Observe that NGF augmented the kinase activity, and this increase was prevented by the presence of DMAT, highlighting the specificity of NGF-induced CK2 activity. (B) NGF activation of CK2 is mediated by p75NTR. Cultured neurons were incubated for 30 min either in the presence or absence of 100 ng/ml NGF or of NGF plus anti-p75-antibody blocking antibody. Note that there is a significant increase of CK2 activity upon exposure of the cells to NGF, and this effect is neutralized by the presence of the antibody. Also note that the inclusion of a rabbit neutral serum did not block NGF activity. (C) Inhibition of CK2 by DMAT prevented NGF-promoted axon elongation. (D) NGF-induced CK2 activation correlates with the NGF-induced phosphorylation of PTEN and Ser9 phosphorylation of GSK-3β. Staining for β3-tubulin was included as a loading control. Observe that NGF did not increase the Ser9 phosphorylation of GSK-3β when CK2 was inhibited by DMAT. (E) Quantification of the densitometric signals from D are the average of three independent experiments.