Huwe1-Mediated Ubiquitylation of Dishevelled Defines a Negative Feedback Loop in the Wnt Signaling Pathway

Abstract

Wnt signaling plays a central role in development, adult tissue homeostasis, and cancer. Several steps in the canonical Wnt/β-catenin signaling cascade are regulated by ubiquitylation, a protein modification that influences the stability, subcellular localization, or interactions of target proteins. To identify regulators of the Wnt/β-catenin pathway, we performed an RNA interference screen in Caenorhabditis elegans and identified the HECT domain-containing ubiquitin ligase EEL-1 as an inhibitor of Wnt signaling. In human embryonic kidney 293T cells, knockdown of the EEL-1 homolog Huwe1 enhanced the activity of a Wnt reporter in cells stimulated with Wnt3a or in cells that overexpressed casein kinase 1 (CK1) or a constitutively active mutant of the Wnt co-receptor low-density lipoprotein receptor-related protein 6 (LRP6). However, knockdown of Huwe1 had no effect on reporter gene expression in cells expressing constitutively active β-catenin, suggesting that Huwe1 inhibited Wnt signaling upstream of β-catenin and downstream of CK1 and LRP6. Huwe1 bound to and ubiquitylated the cytoplasmic Wnt pathway component Dishevelled (Dvl) in a Wnt3a- and CK1ε-dependent manner. Mass spectrometric analysis showed that Huwe1 promoted K63-linked, but not K48-linked, polyubiquitination of Dvl. Instead of targeting Dvl for degradation, ubiquitylation of the DIX domain of Dvl by Huwe1 inhibited Dvl multimerization, which is necessary for its function. Our findings indicate that Huwe1 is part of an evolutionarily conserved negative feedback loop in the Wnt/β-catenin pathway.

Destabilization of Atoh1 by E3 Ubiquitin Ligase Huwe1 and Casein Kinase 1 Is Essential for Normal Sensory Hair Cell Development

“…The regulation of transcription factor expression for the expansion of neural progenitors has been demonstrated in the cerebellum (37), and indeed, Huwe1 regulates the expansion of cerebellar neural progenitors through its action on Myc and ligands of Notch (38). Huwe1 promotes interaction of c-Myc with P300 and transcriptional activation via K63 ubiquitylation (39) and inhibits multimerization of the cytoplasmic scaffolding protein Dvl-3, resulting in down-regulation of Wnt signaling (40). Huwe1 was recently shown to play a role in Atoh1 stability in the cerebellum through which it influenced the differentiation and distribution of cerebellar neural progenitors (15).…”

HUWE1 Is a Molecular Link Controlling RAF-1 Activity Supported by the Shoc2 Scaffold

“…of Shoc2, HUWE1 inhibited the ability of Shoc2 to accelerate ERK1/2 phosphorylation. Though HUWE1 has been shown to synthesize K48 chains on several of its substrates, including p53, ARF, Mcl-1, and N-Myc (46,62,63), it also preferentially assembles K63 chains on c-Myc (52) and on its recently identified substrate Dvl (64). In both cases, HUWE1-induced ubiquitination was shown to be a part of a negative-feedback loop.…”

Potential signaling pathways as therapeutic targets for overcoming chemoresistance in mucinous ovarian cancer (Review)

“…The organic molecule NSC668036 that binds to and blocks the PDZ domain of Dvl also inhibits WNT signaling (54). As alternative methods of inhibition, HUWE1 ubiquitylates Dvl to promote its degradation (55); while WWOX, a member of the short-chain dehydrogenases/reductases protein family, prevents nuclear import of Dvl proteins (56). Similar to HUWE1, DACT1 (also known as dapper antagonist of catenin-1) antagonizes WNT/β-catenin signaling by inducing Dvl degradation (54,57).…”