Axin serves as a scaffold protein in Wnt signaling pathway to negatively regulate the stability of β-catenin, and it also facilitates the c-Jun N-terminal/stress-activated protein kinase (JNK/SAPK) pathway through MEKK1. Several other elements are required to activate JNK, such as homodimerization, sumoylation at the extreme C-terminal sites, and a region in the PP2A binding domain. PP2A binding domain of Axin was suggested to play an important role in facilitating JNK activation. MEKK1 binding Deficiency mutant was made, but it still can activate JNK. We showed that another MEKK family member MEKK4 is a mediator of Axin/JNK activation through binding on PP2A domain of Axin, which is distinct from MEKK1 binding site. It is suggesting that Axin may receive distinct signals tethered by ME...[ Read more ]

Axin serves as a scaffold protein in Wnt signaling pathway to negatively regulate the stability of β-catenin, and it also facilitates the c-Jun N-terminal/stress-activated protein kinase (JNK/SAPK) pathway through MEKK1. Several other elements are required to activate JNK, such as homodimerization, sumoylation at the extreme C-terminal sites, and a region in the PP2A binding domain. PP2A binding domain of Axin was suggested to play an important role in facilitating JNK activation. MEKK1 binding Deficiency mutant was made, but it still can activate JNK. We showed that another MEKK family member MEKK4 is a mediator of Axin/JNK activation through binding on PP2A domain of Axin, which is distinct from MEKK1 binding site. It is suggesting that Axin may receive distinct signals tethered by MEKK1 and MEKK4. Kinase dead mutant of MEKK4 can attenuates the level of JNK activation by MEKK1 binding defective mutant of Axin. We also identified an additional domain between MEKK1 and MEKK4 binding sites as being required for both MEKK1 and MEKK4 to activate JNK via Axin. MEKK1-/- cell line and RNA interference were employed to confirm MEKK4 is indeed a MAP3K that is involved in Axin mediated JNK activation.

On the other hand, by yeast two hybrid screening approach, we used PP2A domain (507-663aa) of Axin as bait and we discover a novel Axin binding protein, p53 ubiquitin ligase (Pirh2). Pirh2 is a negative regulator of p53 activity by inducing p53 degradation. Axin was found to co-immunoprecipitate with Pirh2 and increase transcription activity of p53. Here we showed that Axin might destabilize Pirh2 to facilitate the transcription activity of p53. By ubiquitination assay of Pirh2, ubiquitin signal of Pirh2 was found to increase when Axin was present. Taken together, Axin may induce Pirh2 ubiquitination to destabilize Pirh2 and increase p53 activity. PP2A domain of Axin showed to have importance function in regulating various signaling pathways.