👤 Tor Espen Thorvaldsen

Inhibition of the tankyrase enzymes (TNKS1 and TNKS2) has recently been shown to induce highly dynamic assemblies of β-catenin destruction complex components known as degradasomes, which promote degradation of β-catenin and reduced Wnt signaling activity in colorectal cancer cells. AXIN1 and AXIN2/Conductin, the rate-limiting factors for the stability and function of endogenous destruction complexes, are stabilized upon TNKS inhibition due to abrogated degradation of AXIN by the proteasome. Since the role of AXIN1 versus AXIN2 as scaffolding proteins in the Wnt signaling pathway still remains incompletely understood, we sought to elucidate their relative contribution in the formation of degradasomes, as these protein assemblies most likely represent the morphological and functional correlates of endogenous β-catenin destruction complexes. In SW480 colorectal cancer cells treated with the tankyrase inhibitor (TNKSi) G007-LK we found that AXIN1 was not required for degradasome formation. In contrast, the formation of degradasomes as well as their capacity to degrade β-catenin were considerably impaired in G007-LK-treated cells depleted of AXIN2. These findings give novel insights into differential functional roles of AXIN1 versus AXIN2 in the β-catenin destruction complex. Show less

In canonical Wnt signaling, the protein levels of the key signaling mediator β-catenin are under tight regulation by the multimeric destruction complex that mediates proteasomal degradation of β-catenin. In colorectal cancer, destruction complex activity is often compromised due to mutations in the multifunctional scaffolding protein Adenomatous Polyposis Coli (APC), leading to a stabilization of β-catenin. Recently, tankyrase inhibitors (TNKSi), a novel class of small molecule inhibitors, were shown to re-establish a functional destruction complex in APC-mutant cancer cell lines by stabilizing AXIN1/2, whose protein levels are usually kept low via poly(ADP-ribosyl)ation by the tankyrase enzymes (TNKS1/2). Surprisingly, we found that for the formation of the morphological correlates of destruction complexes, called degradasomes, functional proteasomes are required. In addition we found that AXIN2 is strongly upregulated after 6 h of TNKS inhibition. The proteasome inhibitor MG132 counteracted TNKSi-induced degradasome formation and AXIN2 stabilization, and this was accompanied by reduced transcription of AXIN2. Mechanistically we could implicate the transcription factor FoxM1 in this process, which was recently shown to be a transcriptional activator of AXIN2. We observed a substantial reduction in TNKSi-induced stabilization of AXIN2 after siRNA-mediated depletion of FoxM1 and found that proteasome inhibition reduced the active (phosphorylated) fraction of FoxM1. This can explain the decreased protein levels of AXIN2 after MG132 treatment. Our findings have implications for the design of in vitro studies on the destruction complex and for clinical applications of TNKSi. Show less