TGF-β signaling involves a wide array of signaling molecules and multiple controlling events. Scaffold proteins create a functional proximity of signaling molecules and control the specificity of signal transduction. While many components involved in the TGF-β pathway have been elucidated, little is known about how those components are coordinated by scaffold proteins. Here, we show that Axin, originally identified as a central scaffold to coordinate the degradation of β-catenin in Wnt signaling, activates TGF-β signaling by forming a multimeric complex consisting of Smad7 and ubiquitin E3 ligase Arkadia. Axin depends on Arkadia to facilitate TGF-β signaling, as their siRNAs reciprocally abolished the stimulatory effect on TGF-β signaling. Specific knockdown of Axin or Arkadia indicates...[ Read more ]

TGF-β signaling involves a wide array of signaling molecules and multiple controlling events. Scaffold proteins create a functional proximity of signaling molecules and control the specificity of signal transduction. While many components involved in the TGF-β pathway have been elucidated, little is known about how those components are coordinated by scaffold proteins. Here, we show that Axin, originally identified as a central scaffold to coordinate the degradation of β-catenin in Wnt signaling, activates TGF-β signaling by forming a multimeric complex consisting of Smad7 and ubiquitin E3 ligase Arkadia. Axin depends on Arkadia to facilitate TGF-β signaling, as their siRNAs reciprocally abolished the stimulatory effect on TGF-β signaling. Specific knockdown of Axin or Arkadia indicates that Axin and Arkadia cooperate with each other in promoting Smad7 ubiquitination. Pulse-chase experiments further illustrated that Axin significantly decreased the half-life of Smad7. Axin also induces nuclear export of Smad7. Interestingly, Axin appears to be constitutively associated with Arkadia and Smad7, in contrast to its transient association with inactive Smad3, underscoring its importance as an intrinsic regulator in TGF-β signaling.

In parallel, we have also been embarking on characterization of Axin proteins in the Axin^Fu mice whose phenotype is kinked tails. The Axin^Fu allele is caused by insertion of an intracistemal particle (IAP) that could potentially generate mutant Axin transcripts as well as wildtype Axin. In the present study, using specific antibodies against different regions of Axin, we conducted immunoprecipitation of total brain extracts of the Axin^Fu mice and found that a truncated Axin containing aa 1-596 (designated as Axin^Fu-NT) is present in the brain of mutant mice. When tested for functionality changes, Axin^Fu-NT was found to act as a dominant negative factor in Axin-mediated activation of JNK and p53. It competes with Axin for MEKK1 and MEKK4 binding which are necessary for Axin-induced JNK activation, while the inhibition of p53 signaling is due to defects in cooperating with HIPK2. Furthermore, Axin^Fu-NT effectively downregulates Wnt signaling, but it exhibits no significant effects on Axin-induced TGF-β signaling. Thus, Axin^Fu-NT may cause the kinked tail phenotype by blocking proper signaling of Axin-mediated JNK activation and p53 activation.