Wnt Signaling Requires Retromer-Dependent Recycling of MIG-14/Wntless in Wnt-Producing Cells

Hubrecht Institute, Developmental Biology and Stem Cell Research, Uppsalalaan 8, 3584 CT, Utrecht, The Netherlands.
Developmental Cell (Impact Factor: 9.71). 02/2008; 14(1):140-7. DOI: 10.1016/j.devcel.2007.12.004
Source: PubMed


Wnt proteins are secreted signaling molecules that play a central role in development and adult tissue homeostasis. We have previously shown that Wnt signaling requires retromer function in Wnt-producing cells. The retromer is a multiprotein complex that mediates endosome-to-Golgi transport of specific sorting receptors. MIG-14/Wls is a conserved transmembrane protein that binds Wnt and is required in Wnt-producing cells for Wnt secretion. Here, we demonstrate that in the absence of retromer function, MIG-14/Wls is degraded in lysosomes and becomes limiting for Wnt signaling. We show that retromer-dependent recycling of MIG-14/Wls is part of a trafficking pathway that retrieves MIG-14/Wls from the plasma membrane. We propose that MIG-14/Wls cycles between the Golgi and the plasma membrane to mediate Wnt secretion. Regulation of this transport pathway may enable Wnt-producing cells to control the range of Wnt signaling in the tissue.

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Available from: Magdalena Lorenowicz, May 19, 2015
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    • "In our studies of Wnt secretion, we generated monoclonal antibody YJ5 that recognizes the first predicted extracellular loop of human WLS and showed that this antibody specifically recognizes both overexpressed and endogenous WLS by immunoblotting in multiple cell lines (Coombs et al., 2010). Using YJ5, we confirmed previous reports that WLS with either a V5 or GFP tag at the carboxyl terminus localizes predominantly to the Golgi (Figures 1C–1E; Figure S1A available online) (Yang et al., 2008). However, when we removed the carboxy-terminal V5 epitope tag by the simple expedient of reintroducing the native stop codon, we found that under identical transfection conditions the subcellular localization changed. "
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    ABSTRACT: Wnts are transported to the cell surface by the integral membrane protein WLS (also known as Wntless, Evi, and GPR177). Previous studies of WLS trafficking have emphasized WLS movement from the Golgi to the plasma membrane (PM) and then back to the Golgi via retromer-mediated endocytic recycling. We find that endogenous WLS binds Wnts in the endoplasmic reticulum (ER), cycles to the PM, and then returns to the ER through the Golgi. We identify an ER-targeting sequence at the carboxyl terminus of native WLS that is critical for ER retrograde recycling and contributes to Wnt secretory function. Golgi-to-ER recycling of WLS requires the COPI regulator ARF as well as ERGIC2, an ER-Golgi intermediate compartment protein that is also required for the retrograde trafficking of the KDEL receptor and certain toxins. ERGIC2 is required for efficient Wnt secretion. ER retrieval is an integral part of the WLS transport cycle.
    Developmental Cell 04/2014; 29(3). DOI:10.1016/j.devcel.2014.03.016 · 9.71 Impact Factor
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    • "Importantly, studies in C. elegans, Drosophila and mammalian tissue-culture cells have shown that Wls needs to be retrieved back to the trans-Golgi network (TGN) to maintain Wnt secretion. This retrieval route involves AP-2 and clathrin mediated endocytosis of Wls from the plasma membrane [14–16] and transport from endosomes to the TGN, a retrograde trafficking step that is mediated by the retromer complex [14,15,17–20]. In the absence of a functional retromer complex, Wls is retained in the endosomal system and degraded in lysosomes. "
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    ABSTRACT: In C. elegans and Drosophila, retromer mediated retrograde transport of Wntless (Wls) from endosomes to the trans-Golgi network (TGN) is required for Wnt secretion. When this retrograde transport pathway is blocked, Wls is missorted to lysosomes and degraded, resulting in reduced Wnt secretion and various Wnt related phenotypes. In the mammalian intestine, Wnt signaling is essential to maintain stem cells. This prompted us to ask if retromer mediated Wls recycling is also important for Wnt signaling and stem cell maintenance in this system. To answer this question, we generated a conditional Vps35 (fl) allele. As Vps35 is an essential subunit of the retromer complex, this genetic tool allowed us to inducibly interfere with retromer function in the intestinal epithelium. Using a pan-intestinal epithelial Cre line (Villin-CreERT2), we did not observe defects in crypt or villus morphology after deletion of Vps35 from the intestinal epithelium. Wnt secreted from the mesenchyme of the intestine may compensate for a reduction in epithelial Wnt secretion. To exclude the effect of the mesenchyme, we generated intestinal organoid cultures. Loss of Vps35 in intestinal organoids did not affect the overall morphology of the organoids. We were able to culture Vps35 (∆/∆) organoids for many passages without Wnt supplementation in the growth medium. However, Wls protein levels were reduced and we observed a subtle growth defect in the Vps35 (∆/∆) organoids. These results confirm the role of retromer in the retrograde trafficking of Wls in the intestine, but show that retromer mediated Wls recycling is not essential to maintain Wnt signaling or stem cell proliferation in the intestinal epithelium.
    PLoS ONE 10/2013; 8(10):e76971. DOI:10.1371/journal.pone.0076971 · 3.23 Impact Factor
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    • "The membrane deformation sub-complex consists of sorting nexin (SNX) dimers (Vps5p and Vps17p in yeast, and sortin nexins 1/3 or 5/6 in vertebrates) (van Weering et al., 2010; Seaman, 2005; Bonifacino and Hurley, 2008). A growing list of retromer cargos has been identified, including cation independent mannose 6-phosphate receptors (CI-MRP) (Arighi et al., 2004), wntless (a “receptor” for Wnt morphogens) (Belenkaya et al., 2008; Eaton, 2008; Franch-Marro et al., 2008; Pan et al., 2008; Port et al., 2008; Yang et al., 2008), Ced1 (a phagocytic receptor) (Chen et al., 2010), VPS10/sotilin family proteins, such as VPS10 in yeast (Iwaki et al., 2006), sortilin and sortilin-related receptor (SorL1 or SorCS1) in vertebrates (Canuel et al., 2008; Kim et al., 2010; Okada et al., 2010), and G-protein coupled receptors (e.g., Parathyroid hormone receptor and β2-adrenonergic receptor) (Feinstein et al., 2011; Temkin et al., 2011). Although retromer's endosome-to-Golgi retrieval function has been well established in yeast, C. elegans, Drosophila, and in mammalian cultured cells, its role in mouse just began to be understood. "
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    ABSTRACT: VPS35, a major component of the retromer, plays an important role in the selective endosome-to-Golgi retrieval of membrane proteins. Dysfunction of retromer is a risk factor for neurodegenerative disorders, but its function in developing mouse brain remains poorly understood. Here we provide evidence for VPS35 promoting dendritic growth and maturation, and axonal protein transport in developing mouse hippocampal neurons. Embryonic hippocampal CA1 neurons suppressing Vps35 expression by in utero electroporation of its micro RNAs displayed shortened apical dendrites, reduced dendritic spines, and swollen commissural axons in the neonatal stage, those deficits reflecting a defective protein transport/trafficking in developing mouse neurons. Further mechanistic studies showed that Vps35 depletion in neurons resulted in an impaired retrograde trafficking of BACE1 (β1-secretase) and altered BACE1 distribution. Suppression of BACE1 expression in CA1 neurons partially rescued both dendritic and axonal deficits induced by Vps35-deficiency. These results thus demonstrate that BACE1 acts as a critical cargo of retromer in vitro and in vivo, and suggest that VPS35 plays an essential role in regulating apical dendritic maturation and in preventing axonal spheroid formation in developing hippocampal neurons.
    Biology Open 12/2012; 1(12):1248-57. DOI:10.1242/bio.20122451 · 2.42 Impact Factor
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