Article

Phosphorylation of Dishevelled by Protein Kinase RIPK4 Regulates Wnt Signaling.

Department of Physiological Chemistry, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA.
Science (Impact Factor: 31.48). 01/2013; DOI: 10.1126/science.1232253
Source: PubMed

ABSTRACT Receptor-interacting protein kinase 4 (RIPK4) is required for epidermal differentiation and is mutated in Bartsocas-Papas syndrome. RIPK4 binds to protein kinase C, but its signaling mechanisms are largely unknown. Ectopic RIPK4, but not catalytically inactive or Bartsocas-Papas RIPK4 mutants, induced accumulation of cytosolic β-catenin and a transcriptional program similar to that caused by Wnt3a. In Xenopus embryos, Ripk4 synergized with coexpressed Xwnt8, whereas Ripk4 morpholinos or catalytic inactive Ripk4 antagonized Wnt signaling. RIPK4 interacted constitutively with the adaptor protein DVL2 and, after Wnt3a stimulation, with the coreceptor LRP6. Phosphorylation of DVL2 by RIPK4 favored canonical Wnt signaling. Wnt-dependent growth of xenografted human tumor cells was suppressed by RIPK4 knockdown, suggesting that RIPK4 overexpression may contribute to the growth of certain tumor types.

Download full-text

Full-text

Available from: Vishva Dixit, Jun 21, 2015
2 Followers
 · 
302 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The phosphoprotein Dishevelled (Dvl) is a common essential component of Wnt/β-catenin and Wnt/planar cell polarity (PCP) signaling pathways. However, the regulation and significance of Dvl phosphorylation are not fully understood. Here, we show that homeodomain-interacting protein kinase 2 (Hipk2) facilitates protein phosphatase 1 catalytic subunit (PP1c)-mediated dephosphorylation of Dvl via its C-terminal domain and that this dephosphorylation blocks ubiquitination and consequent degradation mediated by the E3 ubiquitin ligase Itch, which targets the phosphorylated form of Dvl proteins. Inhibition of Hipk2 or PP1c function reduces Dvl protein levels and suppresses Wnt/β-catenin and Wnt/PCP pathway-dependent events in mammalian cells and zebrafish embryos, suggesting that Hipk2 and PP1c are essential for maintaining Dvl protein levels that are sufficient to activate Wnt signaling. We also show that Wnt-3a, a Wnt/β-catenin ligand, induces dissociation of the Dvl-Hipk2-PP1c complex and Dvl degradation under high-cell-density conditions. This regulation may be a negative feedback mechanism that fine-tunes Wnt/β-catenin signaling.
    Cell Reports 09/2014; 8(5). DOI:10.1016/j.celrep.2014.07.040 · 7.21 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Receptor-interacting protein kinase 4 (RIPK4)-deficient mice have epidermal defects and fusion of all external orifices. These are similar to Bartsocas-Papas syndrome and popliteal pterygium syndrome (PPS) in humans, for which causative mutations have been documented in the RIPK4 and IRF6 (interferon regulatory factor 6) gene, respectively. Although genetically distinct, these syndromes share the anomalies of marked pterygia, syndactyly, clefting and hypoplastic genitalia. Despite the strong resemblance of these two syndromes, no molecular connection between the transcription factor IRF6 and the kinase RIPK4 was known and the mechanism underlying the phenotype was unclear. Here we describe that RIPK4 deficiency in mice causes epithelial fusions associated with abnormal periderm development and aberrant ectopic localization of E-cadherin on the apical membrane of the outer peridermal cell layers. In Xenopus, RIPK4 depletion causes the absence of ectodermal epiboly and concomitant gastrulation defects that phenocopy ectopic expression of dominant-negative IRF6. We found that IRF6 controls RIPK4 expression and that wild-type, but not kinase-dead, RIPK4 can complement the gastrulation defect in Xenopus caused by IRF6 malfunctioning. In contrast to the mouse, we observed only minor effects on cadherin membrane expression in Xenopus RIPK4 morphants. However, gastrulation defects were associated with a virtual absence of cortical actin in the ectodermal cells that face the blastocoel cavity and this was phenocopied in embryos expressing dominant-negative IRF6. A role for RIPK4 in actin cytoskeleton organization was also revealed in mouse epidermis and in human epithelial HaCaT cells. In conclusion, we showed that in mice RIPK4 is implicated in cortical actin organization and in E-cadherin localization or function, which can explain the characteristic epithelial fusions observed in PPSs. In addition, we provide a novel molecular link between IRF6 and RIPK4 that unifies the different PPSs to a common molecular pathway.Cell Death and Differentiation advance online publication, 28 November 2014; doi:10.1038/cdd.2014.191.
    Cell Death and Differentiation 11/2014; 22(6). DOI:10.1038/cdd.2014.191 · 8.39 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Receptor-Interacting Protein Kinase 4 (RIPK4) and Interferon Regulatory Factor 6 (IRF6) are critical regulators of keratinocyte differentiation and their mutation cause the related developmental epidermal disorders Bartsocas-Papas Syndrome and Popliteal Pterygium Syndrome, respectively. However, the signaling pathways in which RIPK4 and IRF6 operate to regulate keratinocyte differentiation are poorly defined. Here, we identify and mechanistically define a direct functional relationship between RIPK4 and IRF6. Gene promoter reporter and in vitro kinase assays, co-immunoprecipitation experiments, and confocal microscopy demonstrated that RIPK4 directly regulates IRF6 trans-activator activity and nuclear translocation. Gene knock-down and overexpression studies indicated that the RIPK4-IRF6 signaling axis controls the expression of key transcriptional regulators of keratinocyte differentiation, including Grainyhead-Like 3 and OVO-Like 1. Additionally, we demonstrate that the p.Ile121Asn missense mutation in RIPK4, which was recently identified in Bartsocas-Papas Syndrome, inhibits its kinase activity, thereby preventing RIPK4-mediated IRF6 activation and nuclear translocation. We show through mutagenesis-based experiments that Ser-413 and Ser-424 in IRF6 are important for its activation by RIPK4. RIPK4 is also important for the regulation of IRF6 expression by the protein kinase C pathway. Our findings therefore not only provide important mechanistic insights into the regulation of keratinocyte differentiation by RIPK4 and IRF6, but they also suggest one mechanism by which mutations in RIPK4 may cause epidermal disorders (e.g. Bartsocas-Papas Syndrome), namely by the impaired activation of IRF6 by RIPK4.
    Journal of Biological Chemistry 09/2014; DOI:10.1074/jbc.M114.589382 · 4.60 Impact Factor