Cytosolic accumulation of γH2AX is associated with tropomyosin-related kinase A-induced cell death in U2OS cells

3-kinase; Tet, tetracycline; TrkA, tropomyosin-related kinase A Tropomyosin-related kinase A (TrkA) plays an im-portant role in cell survival, differentiation, and apopto-sis in various neuronal and nonneuronal cell types. Here we show that TrkA overexpression by the Tet-On system mimics NGF-mediated activation pathways in the absence of nerve growth factor (NGF) stimulation in U2OS cells. In addition, p53 upregulation upon DNA damage was inhibited by TrkA, and p21 was upregu-lated by TrkA in a p53-independent manner. TrkA over-expression caused cell death by interrupting cell cycle progression, and TrkA-induced cell death was dimin-ished in the presence of its specific inhibitor GW441756. Interestingly, TrkA-mediated cell death was strongly related to γH2AX production and poly (ADP-ribose) polymerase cleavage in the absence of DNA damage inducer. In this study, we also reveal that γH2AX production by TrkA is blocked by TrkA kinase inhibitors K-252a and GW441756, and it is also sig-nificantly inhibited by JNK inhibitor SP600125. More-over, reduction of cell viability by TrkA was strongly suppressed by SP600125 treatment, suggesting a crit-ical role of JNK in TrkA-induced cell death. We also found that γH2AX and TrkA were colocalized in cytosol in the absence of DNA damage, and the nuclear local-ization of γH2AX induced by DNA damage was partly altered to cytosol by TrkA overexpression. Our results suggest that the abnormal cytosolic accumulation of γH2AX is implicated in TrkA-induced cell death in the absence of DNA damage