Ape1/Ref-1 Stimulates GDNF/GFRalpha1-mediated Downstream Signaling and Neuroblastoma Proliferation

We previously reported that glial cell line-derived neurotropic factor (GDNF) receptor alpha1 (GFRalpha1) is a direct target of apurinic/apyrimidinic endonuclease 1 (Ape1/Ref-1). In the present study, we further analyzed the physiological roles of Ape1/Ref-1-induced GFRalpha1 expression in Neuro2a mouse neuroblastoma cells. Ape1/Ref-1 expression caused the clustering of GFRalpha1 immunoreactivity in lipid rafts in response to GDNF. We also found that Ret, a downstream target of GFRalpha1, was functionally activated by GDNF in Ape1/Ref-1-expressing cells. Moreover, GDNF promoted the proliferation of Ape1/Ref-1-expressing Neuro2a cells. Furthermore, GFRalpha1-specific RNA experiments demonstrated that the downregulation of GFRalpha1 by siRNA in Ape1/Ref-1-expressing cells impaired the ability of GDNF to phosphorylate Akt and PLCgamma-1 and to stimulate cellular proliferation. These results show an association between Ape1/Ref-1 and GDNF/GFRalpha signaling, and suggest a potential molecular mechanism for the involvement of Ape1/Ref-1 in neuronal proliferation.