title: Gtpbp2 is a positive regulator of Wnt signaling and maintains low levels of the Wnt negative regulator Axin.
abstract: Canonical Wnt signals, transduced by stabilized β-catenin, play similar roles across animals in maintaining stem cell pluripotency, regulating cell differentiation, and instructing normal embryonic development. Dysregulated Wnt/β-catenin signaling causes diseases and birth defects, and a variety of regulatory processes control this pathway to ensure its proper function and integration with other signaling systems. We previously identified GTP-binding protein 2 (Gtpbp2) as a novel regulator of BMP signaling, however further exploration revealed that Gtpbp2 can also affect Wnt signaling, which is a novel finding reported here.
  
  
  
  Knockdown of Gtpbp2 in Xenopus embryos causes severe axial defects and reduces expression of Spemann-Mangold organizer genes. Gtpbp2 knockdown blocks responses to ectopic Wnt8 ligand, such as organizer gene induction in ectodermal tissue explants and induction of secondary axes in whole embryos. However, organizer gene induction by ectopic Nodal2 is unaffected by Gtpbp2 knockdown. Epistasis tests, conducted by activating Wnt signal transduction at sequential points in the canonical pathway, demonstrate that Gtpbp2 is required downstream of Dishevelled and Gsk3β but upstream of β-catenin, which is similar to the previously reported effects of Axin1 overexpression in Xenopus embryos. Focusing on Axin in Xenopus embryos, we find that knockdown of Gtpbp2 elevates endogenous or exogenous Axin protein levels. Furthermore, Gtpbp2 fusion proteins co-localize with Dishevelled and co-immunoprecipitate with Axin and Gsk3b.
  
  
  
  We conclude that Gtpbp2 is required for canonical Wnt/β-catenin signaling in Xenopus embryos. Our data suggest a model in which Gtpbp2 suppresses the accumulation of Axin protein, a rate-limiting component of the β-catenin destruction complex, such that Axin protein levels negatively correlate with Gtpbp2 levels. This model is supported by the similarity of our Gtpbp2-Wnt epistasis results and previously reported effects of Axin overexpression, the physical interactions of Gtpbp2 with Axin, and the correlation between elevated Axin protein levels and lost Wnt responsiveness upon Gtpbp2 knockdown. A wide variety of cancer-causing Wnt pathway mutations require low Axin levels, so development of Gtpbp2 inhibitors may provide a new therapeutic strategy to elevate Axin and suppress aberrant β-catenin signaling in cancer and other Wnt-related diseases.
authors.affiliation: Department of Biochemistry and Cell Biology, Graduate Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY, 11794-5215, USA.|Present Address: Department of Biological Sciences, State University of New York, College at Old Westbury, Old Westbury, NY, 11568, USA.
authors.forename: William Q
authors.initials: WQ
authors.lastname: Gillis
authors.affiliation: Department of Biochemistry and Cell Biology, Graduate Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY, 11794-5215, USA.|Present Address: The Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Headington, Oxford, OX3 9DS, UK.
authors.forename: Arif
authors.initials: A
authors.lastname: Kirmizitas
authors.affiliation: Department of Biochemistry and Cell Biology, Graduate Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY, 11794-5215, USA.
authors.forename: Yasuno
authors.initials: Y
authors.lastname: Iwasaki
authors.affiliation: Department of Biochemistry and Cell Biology, Graduate Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY, 11794-5215, USA.|Present Address: Department of Pediatric Oncology, Dana-Farber Cancer Institute, Division of Hematology/Oncology, Children's Hospital Boston, Harvard Medical School, Boston, MA, USA.
authors.forename: Dong-Hyuk
authors.initials: DH
authors.lastname: Ki
authors.affiliation: Department of Biochemistry and Cell Biology, Graduate Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY, 11794-5215, USA.
authors.forename: Jonathan M
authors.initials: JM
authors.lastname: Wyrick
authors.affiliation: Department of Biochemistry and Cell Biology, Graduate Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY, 11794-5215, USA. gerald.h.thomsen@stonybrook.edu.
authors.forename: Gerald H
authors.initials: GH
authors.lastname: Thomsen
mesh_terms: D000818:Animals; D060466:Axin Protein / Q000235:genetics / Q000378:metabolism*; D000072261:Dishevelled Proteins / Q000378:metabolism; D004843:Epistasis, Genetic; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism; D011485:Protein Binding; D060449:Wnt Signaling Pathway; D014981:Xenopus; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D060466:Axin Protein; D000072261:Dishevelled Proteins; D029867:Xenopus Proteins; D000071679:Glycogen Synthase Kinase 3 beta; D020558:GTP Phosphohydrolases; C000591592:Gtpbp2 protein, Xenopus
keywords: Axin; GTPase; Gtpbp2; Wnt signaling; Xenopus embryo