Arkadia, a novel SUMO-targeted ubiquitin ligase involved in PML degradation.

Arkadia is a RING domain E3 ubiquitin ligase that activates the transforming growth factor ß (TGF-ß) pathway by inducing degradation of the inhibitor SnoN/Ski. Here we show that Arkadia contains three successive SUMO-interacting motifs (SIMs) that mediate noncovalent interaction with poly-SUMO2. We identify the third SIM (VVDL) of Arkadia to be the most relevant one in this interaction. Furthermore, we provide evidence that Arkadia can function as a SUMO-targeted ubiquitin ligase (STUBL) by ubiquitinating SUMO chains. While the SIMs of Arkadia are not essential for SnoN/Ski degradation in response to TGF-ß, we show that they are necessary for the interaction of Arkadia with polysumoylated PML in response to arsenic and its concomitant accumulation into PML nuclear bodies. Moreover, Arkadia depletion leads to accumulation of polysumoylated PML in response to arsenic, highlighting a requirement of Arkadia for arsenic-induced degradation of polysumoylated PML. Interestingly, Arkadia homodimerizes but does not heterodimerize with RNF4, the other STUBL involved in PML degradation, suggesting that these two E3 ligases do not act synergistically but most probably act independently during this process. Altogether, these results identify Arkadia to be a novel STUBL that can trigger degradation of signal-induced polysumoylated proteins.

Arkadia regulates tumor metastasis by modulation of the TGF-ß pathway.

TGF-ß can act as a tumor suppressor at early stages of cancer progression and as a tumor promoter at later stages. The E3 ubiquitin ligase Arkadia (RNF111) is a critical component of the TGF-ß signaling pathway, being required for a subset of responses, those mediated by Smad3-Smad4 complexes. It acts by mediating ligand-induced degradation of Ski and SnoN (SKIL), which are 2 potent transcriptional repressors. Here, we investigate the role of Arkadia in cancer using model systems to address both potential tumor-suppressive and tumor-promoting roles. Stable reexpression of Arkadia in lung carcinoma NCI-H460 cells, which we show contain a hemizygous nonsense mutation in the Arkadia/RNF111 gene, efficiently restored TGF-ß-induced Smad3-dependent transcription, and substantially decreased the ability of these cells to grow in soft agar in vitro. However, it had no effect on tumor growth in vivo in mouse models. Moreover, loss of Arkadia in cancer cell lines and human tumors is rare, arguing against a prominent tumor-suppressive role. In contrast, we have uncovered a potent tumor-promoting function for Arkadia. Using 3 different cancer cell lines whose tumorigenic properties are driven by TGF-ß signaling, we show that loss of Arkadia function, either by overexpression of dominant negative Arkadia or by siRNA-induced knockdown, substantially inhibited lung colonization in tail vein injection experiments in immunodeficient mice. Our findings indicate that Arkadia is not critical for regulating tumor growth per se, but is required for the early stages of cancer cell colonization at the sites of metastasis.