Rab8 Regulates Basolateral Secretory, But Not Recycling, Traffic at the Recycling Endosome

Department of Pharmacology, University of Iowa Carver College of Medicine, Iowa City, IA 52242, USA.
Molecular biology of the cell (Impact Factor: 4.47). 06/2008; 19(5):2059-68. DOI: 10.1091/mbc.E07-09-0902
Source: PubMed

ABSTRACT Rab8 is a monomeric GTPase that regulates the delivery of newly synthesized proteins to the basolateral surface in polarized epithelial cells. Recent publications have demonstrated that basolateral proteins interacting with the mu1-B clathrin adapter subunit pass through the recycling endosome (RE) en route from the TGN to the plasma membrane. Because Rab8 interacts with these basolateral proteins, these findings raise the question of whether Rab8 acts before, at, or after the RE. We find that Rab8 overexpression during the formation of polarity in MDCK cells, disrupts polarization of the cell, explaining how Rab8 mutants can disrupt basolateral endocytic and secretory traffic. However, once cells are polarized, Rab8 mutants cause mis-sorting of newly synthesized basolateral proteins such as VSV-G to the apical surface, but do not cause mis-sorting of membrane proteins already at the cell surface or in the endocytic recycling pathway. Enzymatic ablation of the RE also prevents traffic from the TGN from reaching the RE and similarly results in mis-sorting of newly synthesized VSV-G. We conclude that Rab8 regulates biosynthetic traffic through REs to the plasma membrane, but not trafficking of endocytic cargo through the RE. The data are consistent with a model in which Rab8 functions in regulating the delivery of TGN-derived cargo to REs.

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    • "During intestinal differentiation, the intestinal cell fate activator Cdx2 transcriptionally regulates the expression of Rab8 small GTPases, which are members of the Ras superfamily (Gao and Kaestner, 2010). Rab8 directly binds isoforms of the myosin V motor (Hume et al., 2001; Rodriguez and Cheney, 2002; Roland et al., 2009), facilitating exocytotic cargo movements on actin tracks in epithelial and non-epithelial cells (Ang et al., 2003; Bryant et al., 2010; Gerges et al., 2004; Hattula et al., 2006; Henry and Sheff, 2008; Huber et al., 1993a,b; Sato et al., 2009; Sun et al., 2014). Global Rab8a ablation in mice impairs the apical delivery of peptidases and nutrient transporters to enterocyte brush borders; as a consequence, these proteins are transported into lysosomes, causing nutrient deprivation and postnatal death of knockout mice (Sato et al., 2007). "
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    ABSTRACT: Communication between stem and niche supporting cells maintains the homeostasis of adult tissues. Wnt signaling is a crucial regulator of the stem cell niche, but the mechanism that governs Wnt ligand delivery in this compartment has not been fully investigated. We identified that Wnt secretion is partly dependent on Rab8a-mediated anterograde transport of Gpr177 (wntless), a Wnt-specific transmembrane transporter. Gpr177 binds to Rab8a, depletion of which compromises Gpr177 traffic, thereby weakening the secretion of multiple Wnts. Analyses of generic Wnt/β-catenin targets in Rab8a knockout mouse intestinal crypts indicate reduced signaling activities; maturation of Paneth cells - a Wnt-dependent cell type - is severely affected. Rab8a knockout crypts show an expansion of Lgr5(+) and Hopx(+) cells in vivo. However, in vitro, the knockout enteroids exhibit significantly weakened growth that can be partly restored by exogenous Wnts or Gsk3β inhibitors. Immunogold labeling and surface protein isolation identified decreased plasma membrane localization of Gpr177 in Rab8a knockout Paneth cells and fibroblasts. Upon stimulation by exogenous Wnts, Rab8a-deficient cells show ligand-induced Lrp6 phosphorylation and transcriptional reporter activation. Rab8a thus controls Wnt delivery in producing cells and is crucial for Paneth cell maturation. Our data highlight the profound tissue plasticity that occurs in response to stress induced by depletion of a stem cell niche signal. © 2015. Published by The Company of Biologists Ltd.
    Development 05/2015; 128(13). DOI:10.1242/dev.121046 · 6.46 Impact Factor
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    • "Basolateral protein transport from median Golgi and TGN is also carried out by Rab8 and Rab10 [50] [51]. Localization of Rab17 and Rab25 to apical recycling endosomes (APEs) facilitates transcytic transport to the apical and basolateral plasma membranes [52] [32]. "
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    ABSTRACT: Vesicular/Membrane trafficking essentially regulates the compartmentalization and abundance of proteins within the cells and contributes in many signalling pathways. This membrane transport in eukaryotic cells is a complex process regulated by a large and diverse array of proteins. A large group of monomeric small GTPases; the Rabs are essential components of this membrane trafficking route. Most of the Rabs are ubiquitously expressed proteins and have been implicated in vesicle formation, vesicle motility/delivery along cytoskeleton elements and docking/fusion at target membranes through the recruitment of effectors. Functional impairments of Rabs affecting transport pathways manifest different diseases. Rab functions are accompanied by cyclical activation and inactivation of GTP-bound and GDP-bound forms between the cytosol and membranes which is regulated by upstream regulators. Rab proteins are characterized by their distinct sub-cellular localization and regulate a wide variety of endocytic, transcytic and exocytic transport pathways. Mutations of Rabs affect cell growth, motility and other biological processes.
    Experimental Cell Research 07/2014; 328(1). DOI:10.1016/j.yexcr.2014.07.027 · 3.25 Impact Factor
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    • "In addition, Rab proteins colocalize with glial inclusions containing αS in multiple system atrophy (Dalfo and Ferrer, 2005; Nakamura et al., 2000), Rab GTPases were found in vesicle clusters induced by αS overexpression in yeast (Gitler et al., 2008), and overexpression of the Rab GTPases Rab1, Rab3a and Rab8a decreased αS-induced neurotoxicity (Cooper et al., 2006; Gitler et al., 2008). The Rab GTPase Rab8a, which showed the strongest rescue of αS-toxicity in the nematode PD model (Gitler et al., 2008), is associated with recycling endosomes and basolateral trafficking events from the trans-Golgi network to the plasma membrane (Henry and Sheff, 2008). Also, Golgi fragmentation was recently shown to be Rab and SNARE dependent in cellular models of PD, further linking Rab8 with αS (Rendon et al., 2013). "
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    ABSTRACT: Alpha-Synuclein (αS) misfolding is associated with Parkinson's disease (PD) but little is known about the mechanisms underlying αS toxicity. Increasing evidence suggests that defects in membrane transport play an important role in neuronal dysfunction. Here we demonstrate that the GTPase Rab8a interacts with αS in rodent brain. NMR spectroscopy reveals that the C-terminus of αS binds to the functionally important switch region as well as the C-terminal tail of Rab8a. In line with a direct Rab8a/αS interaction, Rab8a enhanced αS aggregation and reduced αS-induced cellular toxicity. In addition, Rab8 - the Drosophila ortholog of Rab8a - ameliorated αS-oligomer specific locomotor impairment and neuron loss in fruit flies. In support of the pathogenic relevance of the αS-Rab8a interaction, phosphorylation of αS at S129 enhanced binding to Rab8a, increased formation of insoluble αS aggregates and reduced cellular toxicity. Our study provides novel mechanistic insights into the interplay of the GTPase Rab8a and αS cytotoxicity, and underscores the therapeutic potential of targeting this interaction.
    Neurobiology of Disease 06/2014; 70. DOI:10.1016/j.nbd.2014.06.018 · 5.08 Impact Factor
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