Elucidation of molecular mechanisms and biological functions of Axin-mediated JNK pathway andp53 signaling.

摘要

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 beta-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.