Elucidation of molecular mechanisms and biological functions of axin-mediated JNK pathway and p53 signaling

Axin is a scaffold protein that controls multiple important pathways, including the canonical Wnt pathway and JNK signaling. We have identified a novel Axin-interacting protein, Aida, which blocks Axin-mediated JNK activation by disrupting Axin homodimerization. During investigation of in-vivo functions of Axin/JNK signaling and aida in development, we have unexpectedly discovered that Axin itself possesses a dorsalizing activity, in addition to ventralizing activity by facilitating β-catenin degradation. This dorsalizing activity is repressed when aida is overexpressed in zebrafish embryo. Whereas Aida-morpholino (Aida-MO) injection led to dorsalized embryos, JNK-MO and MKK4-MO each can ventralize embryos. The anti-dorsalization activity of aida is conferred by its ability to block Axin-mediated JNK activity. We further demonstrate that dorsoventral patterning regulated by Axin/JNK signaling is independent of maternal or zygotic Wnt signaling. We have thus identified a novel dorsalization pathway that is exerted by Axin/JNK signaling and its inhibitor Aida during vertebrate embryogenesis. In parallel, we investigated the functional relationship between Axin, p53 and HIPK2. Axin and p53 are tumor suppressors, controlling cell growth, apoptosis, and development. We show that Axin interacts with homeodomain-interacting protein Kinase-2 (HIPK2), which is linked to UV-induced p53-dependent apoptosis by interacting with, and phosphorylating Ser 46 of, p53. In addition to association with p53 via HIPK2, Axin contains a separate domain that directly interacts with p53 at their physiological concentrations. Axin stimulates p53-dependent reporter transcription in 293 cells, but not in 293T, H1299, or SaOS-2 cells that are defective in p53 signaling. Axin, but not AxindeltaHIPK2, activates HIPK2-mediated p53 phosphorylation at Ser 46, facilitating p53-dependent transcriptional activity and apoptosis. Specific knockdown of Axin by siRNA reduced UV-induced Ser-46 phosphorylation and apoptosis. Kinase-dead HIPK2 reduced Axin-induced p53-dependent transcriptional activity, indicating that Axin stimulates p53 function through HIPK2 kinase activity. Interestingly, HIPK2deltaAxin that lacks its Axin-binding region acts as a dominant-positive form in p53 activation, suggesting that the Axin-binding region of HIPK2 is a putative autoinhibitory domain. These results show that Axin acts as a tumor suppressor by facilitating p53 function through integration of multiple factors