Bighead is a Wnt antagonist secreted by the
            <i>Xenopus</i>
            Spemann organizer that promotes Lrp6 endocytosis

Ding, Yi; Colozza, Gabriele; Sosa, Eric; Moriyama, Yuki; Rundle, Samantha; Salwiński, Łukasz; Robertis, Edward M. De

doi:10.1073/pnas.1812117115

Cited by 39 publications

(40 citation statements)

References 63 publications

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“…We confirmed that Lrp6 endocytosis was observed in SW480, a colon cancer cell line in which the absence of APC triggered, and APC reconstitution inhibited, Wnt signaling and Lrp6 endocytosis (Supplementary Movie S1 ); this is in agreement with recent results showing that APC is a major regulator of endocytosis 16 , 29 . As expected for regulators of Lrp6 endocytosis, the Wnt antagonists Dkk1 30 or Bighead 31 induced endocytosis of Lrp6-APEX2 into large vesicle clusters in the cytoplasm next to the nuclear bay region, where the lysosomal system is located (Fig. 1 K,L).…”

Section: Resultssupporting

confidence: 70%

Wnt-inducible Lrp6-APEX2 interacting proteins identify ESCRT machinery and Trk-fused gene as components of the Wnt signaling pathway

Colozza

Jami‐Alahmadi

Dsouza

et al. 2020

Sci Rep

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The canonical Wnt pathway serves as a hub connecting diverse cellular processes, including β-catenin signaling, differentiation, growth, protein stability, macropinocytosis, and nutrient acquisition in lysosomes. We have proposed that sequestration of β-catenin destruction complex components in multivesicular bodies (MVBs) is required for sustained canonical Wnt signaling. In this study, we investigated the events that follow activation of the canonical Wnt receptor Lrp6 using an APEX2-mediated proximity labeling approach. The Wnt co-receptor Lrp6 was fused to APEX2 and used to biotinylate targets that are recruited near the receptor during Wnt signaling at different time periods. Lrp6 proximity targets were identified by mass spectrometry, and revealed that many endosomal proteins interacted with Lrp6 within 5 min of Wnt3a treatment. Interestingly, we found that Trk-fused gene (TFG), previously known to regulate the cell secretory pathway and to be rearranged in thyroid and lung cancers, was strongly enriched in the proximity of Lrp6. TFG depletion with siRNA, or knock-out with CRISPR/Cas9, significantly reduced Wnt/β-catenin signaling in cell culture. In vivo, studies in the Xenopus system showed that TFG is required for endogenous Wnt-dependent embryonic patterning. The results suggest that the multivesicular endosomal machinery and the novel player TFG have important roles in Wnt signaling.

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Section: Resultssupporting

confidence: 70%

Wnt-inducible Lrp6-APEX2 interacting proteins identify ESCRT machinery and Trk-fused gene as components of the Wnt signaling pathway

Colozza

Jami‐Alahmadi

Dsouza

et al. 2020

Sci Rep

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“…S6E-G), indicating that TFG is required for Wnt activity in vivo. To show that TGF-MO is required for endogenous late Wnt signaling we used knock-down of the Wnt antagonist Bighead, which is known to result in an increase of Wnt activity(31). We found that while Bighead-MO caused loss of anterior structures in Xenopus embryos, this effect could be rescued by co-injection of TFG-MO (Fig.…”

mentioning

confidence: 88%

Wnt-inducible Lrp6-APEX2 Interacting Proteins Identify ESCRT Machinery and Trk-Fused Gene as Components of the Wnt Signaling Pathway

Colozza

Jami‐Alahmadi²,

Dsouza

et al. 2020

Preprint

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The canonical Wnt signaling pathway serves as a hub connecting diverse cellular physiological processes, such as β-catenin signaling, differentiation, growth, protein stability, macropinocytosis, and nutrient acquisition in lysosomes. We have proposed that sequestration of β-catenin destruction complex components in multivesicular bodies (MVBs) is required for sustained canonical Wnt signaling. In this study, we investigated the events that follow activation of the canonical Wnt receptor Lrp6 using an APEX2-mediated proximity labeling approach. The Wnt co-receptor Lrp6 was fused to APEX2 and used to biotinylate targets that are recruited near the receptor during Wnt signaling at different time periods. Lrp6 proximity targets were identified by mass spectrometry, and revealed that many components of the ESCRT (Endocytic Sorting Components Required for Transport) machinery interacted with Lrp6 within 5 minutes of Wnt3a treatment. This supports the proposal of a central role of multivesicular endosomes in canonical Wnt signaling. Interestingly, proteomic analyses identified the Trk-fused gene (TFG), previously known to regulate the cell secretory pathway and to be rearranged in thyroid and lung cancers, as being strongly enriched in the proximity of Lrp6. We provide evidence that TFG specifically co-localized with MVBs after Wnt stimulation. TFG depletion with siRNA, or knock-out with CRISPR/Cas9, significantly reduced Wnt/β-catenin signaling in cell culture. In vivo, studies in the Xenopus system showed that TFG is required for endogenous Wnt-dependent embryonic patterning. The results suggest that the multivesicular endosomal machinery and the novel player TFG have important roles in Wnt signaling. SignificanceWnt/β-catenin signaling is a conserved pathway involved in cell differentiation and in the regulation of many other processes, including cell growth and proliferation, macropinocytosis, and cell metabolism. Endocytosis is required to regulate Wnt signaling, but the precise factors at play are still elusive. Here, we describe a biotin-dependent proximity labeling approach using ascorbate peroxidase-tagged Lrp6, a Wnt co-receptor. Proteomic analysis of biotinylatedenriched targets identified numerous multivesicular endosome proteins that were recruited to the receptor shortly after addition of Wnt protein. Additionally, we identified the protein TFG as one of the strongest interactors with Lrp6. TFG co-localized with Wnt-induced multivesicular endosomes. Xenopus embryo assays revealed that TFG is required in vivo for canonical Wnt signaling. \body

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“…In addition to Dkk1 and Frzb1, other extracellular WNT signaling antagonists such as Bighead, Notum and Tiki1 have been identified in Xenopus embryo studies (Ding et al, 2018;Zhang et al, 2012Zhang et al, , 2015.…”

Section: Molecul Ar Targ E Ts Of Lhx1mentioning

confidence: 99%

Mechanistic insights from the LHX1‐driven molecular network in building the embryonic head

et al. 2019

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Development of an embryo is driven by a series of molecular instructions that control the differentiation of tissue precursor cells and shape the tissues into major body parts. LIM homeobox 1 (LHX1) is a transcription factor that plays a major role in the development of the embryonic head of the mouse. Loss of LHX1 function disrupts the morphogenetic movement of head tissue precursors and impacts on the function of molecular factors in modulating the activity of the WNT signaling pathway. LHX1 acts with a transcription factor complex to regulate the transcription of target genes in multiple phases of development and in a range of embryonic tissues of the mouse and Xenopus. Determining the interacting factors and transcriptional targets of LHX1 will be key to unraveling the ensemble of factors involved in head development and building a head gene regulatory network. K E Y W O R D S gene regulatory network, head development, Lhx1, mouse, Xenopus

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Bighead is a Wnt antagonist secreted by the Xenopus Spemann organizer that promotes Lrp6 endocytosis

Cited by 39 publications

References 63 publications

Wnt-inducible Lrp6-APEX2 interacting proteins identify ESCRT machinery and Trk-fused gene as components of the Wnt signaling pathway

Wnt-inducible Lrp6-APEX2 interacting proteins identify ESCRT machinery and Trk-fused gene as components of the Wnt signaling pathway

Wnt-inducible Lrp6-APEX2 Interacting Proteins Identify ESCRT Machinery and Trk-Fused Gene as Components of the Wnt Signaling Pathway

Mechanistic insights from the LHX1‐driven molecular network in building the embryonic head

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