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Gopinathan K Menon - One of the best experts on this subject based on the ideXlab platform.
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autosomal recessive keratoderma ichthyosis deafness arkid syndrome is caused by vps33b mutations affecting Rab Protein interaction and collagen modification
Journal of Investigative Dermatology, 2017Co-Authors: Robert Gruber, Clare Rogerson, Christian Windpassinger, Blerida Banushi, Anna Straatmaniwanowska, Joanna Hanley, Federico Forneris, Robert Strohal, Debra Crumrine, Gopinathan K MenonAbstract:In this paper, we report three patients with severe palmoplantar keratoderma associated with ichthyosis and sensorineural deafness. Biallelic mutations were found in VPS33B, encoding VPS33B, a Sec1/Munc18 family Protein that interacts with Rab11a and Rab25 Proteins and is involved in trafficking of the collagen-modifying enzyme LH3. Two patients were homozygous for the missense variant p.Gly131Glu, whereas one patient was compound heterozygous for p.Gly131Glu and the splice site mutation c.240-1G>C, previously reported in patients with arthrogryposis renal dysfunction and cholestasis syndrome. We demonstrated the pathogenicity of variant p.Gly131Glu by assessing the interactions of the mutant VPS33B construct and its ability to traffic LH3. Compared with wild-type VPS33B, the p.Gly131Glu mutant VPS33B had reduced coimmunoprecipitation and colocalization with Rab11a and Rab25 and did not rescue LH3 trafficking. Confirming the cell-based experiments, we found deficient LH3-specific collagen lysine modifications in patients’ urine and skin fibroblasts. Additionally, the epidermal ultrastructure of the p.Gly131Glu patients mirrored defects in tamoxifen-inducible VPS33B-deficient Vps33bfl/fl-ERT2 mice. Both patients and murine models revealed an impaired epidermal structure, ascribed to aberrant secretion of lamellar bodies, which are essential for epidermal barrier formation. Our results demonstrate that p.Gly131Glu mutant VPS33B causes an autosomal recessive keratoderma-ichthyosis-deafness syndrome.
Robert Gruber - One of the best experts on this subject based on the ideXlab platform.
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autosomal recessive keratoderma ichthyosis deafness arkid syndrome is caused by vps33b mutations affecting Rab Protein interaction and collagen modification
Journal of Investigative Dermatology, 2017Co-Authors: Robert Gruber, Clare Rogerson, Christian Windpassinger, Blerida Banushi, Anna Straatmaniwanowska, Joanna Hanley, Federico Forneris, Robert Strohal, Debra Crumrine, Gopinathan K MenonAbstract:In this paper, we report three patients with severe palmoplantar keratoderma associated with ichthyosis and sensorineural deafness. Biallelic mutations were found in VPS33B, encoding VPS33B, a Sec1/Munc18 family Protein that interacts with Rab11a and Rab25 Proteins and is involved in trafficking of the collagen-modifying enzyme LH3. Two patients were homozygous for the missense variant p.Gly131Glu, whereas one patient was compound heterozygous for p.Gly131Glu and the splice site mutation c.240-1G>C, previously reported in patients with arthrogryposis renal dysfunction and cholestasis syndrome. We demonstrated the pathogenicity of variant p.Gly131Glu by assessing the interactions of the mutant VPS33B construct and its ability to traffic LH3. Compared with wild-type VPS33B, the p.Gly131Glu mutant VPS33B had reduced coimmunoprecipitation and colocalization with Rab11a and Rab25 and did not rescue LH3 trafficking. Confirming the cell-based experiments, we found deficient LH3-specific collagen lysine modifications in patients’ urine and skin fibroblasts. Additionally, the epidermal ultrastructure of the p.Gly131Glu patients mirrored defects in tamoxifen-inducible VPS33B-deficient Vps33bfl/fl-ERT2 mice. Both patients and murine models revealed an impaired epidermal structure, ascribed to aberrant secretion of lamellar bodies, which are essential for epidermal barrier formation. Our results demonstrate that p.Gly131Glu mutant VPS33B causes an autosomal recessive keratoderma-ichthyosis-deafness syndrome.
Miguel C. Seabra - One of the best experts on this subject based on the ideXlab platform.
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HEMATOPOIESIS A role for Rab27b in NF-E2–dependent pathways of platelet formation
2016Co-Authors: Sanjay Tiwari, Duarte C. Barral, Emilie H. Mules, Miguel C. Seabra, Edward K. Novak, Richard T. Swank, Joseph E. Italiano, Ramesh A. ShivdasaniAbstract:Megakaryocytes release platelets by reor-ganizing the cytoplasm into proplatelet extensions. Fundamental to this process is the need to coordinate transport of products and organelles in the appropri-ate abundance to nascent platelets. The importance of the Rab family of small GTPases (guanosine 5-triphosphatases) in platelet biogenesis is revealed in gun-metal (gm/gm) mice, which show defi-cient Rab isoprenylation and macrothrom-bocytopenia with few granules and abnormal megakaryocyte morphology. Al-though some Rab Proteins are implicated in vesicle and organelle transport along microtubules or actin, the role of any Rab Protein in platelet biogenesis is unknown. The limited number of Rab Proteins with defective membrane association in gm/gm megakaryocytes prominently includes Rab27a and Rab27b. Normal expression of Rab27b is especially increased with terminal megakaryocyte differentiation and dependent on nuclear factor–erythroid 2 (NF-E2), a transcription factor required for thrombopoiesis. Chromatin immuno-precipitation demonstrates recruitment of NF-E2 to the putative Rab27B promoter. Inhibition of endogenous Rab27 function in primary megakaryocytes causes se-vere quantitative and qualitative defects in proplatelet formation that mimic find-ings in gm/gm cells. Rab27b localizes to alpha and dense granules in megakaryo-cytes. These results establish a role for Rab27 in platelet synthesis and suggest that Rab27b in particular may coordinate proplatelet formation with granule trans-port, possibly by recruiting specific effec-tor pathways. (Blood. 2003;102:3970-3979) © 2003 by The American Society of Hematolog
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A role for Rab27b in NF-E2-dependent pathways of platelet formation
Blood, 2003Co-Authors: Sanjay Tiwari, Duarte C. Barral, Emilie H. Mules, Miguel C. Seabra, Edward K. Novak, Richard T. Swank, Joseph E. Italiano, Ramesh A. ShivdasaniAbstract:Megakaryocytes release platelets by reorganizing the cytoplasm into proplatelet extensions. Fundamental to this process is the need to coordinate transport of products and organelles in the appropriate abundance to nascent platelets. The importance of the Rab family of small GTPases (guanosine 5'-triphosphatases) in platelet biogenesis is revealed in gunmetal (gm/gm) mice, which show deficient Rab isoprenylation and macrothrombocytopenia with few granules and abnormal megakaryocyte morphology. Although some Rab Proteins are implicated in vesicle and organelle transport along microtubules or actin, the role of any Rab Protein in platelet biogenesis is unknown. The limited number of Rab Proteins with defective membrane association in gm/gm megakaryocytes prominently includes Rab27a and Rab27b. Normal expression of Rab27b is especially increased with terminal megakaryocyte differentiation and dependent on nuclear factor-erythroid 2 (NF-E2), a transcription factor required for thrombopoiesis. Chromatin immunoprecipitation demonstrates recruitment of NF-E2 to the putative Rab27B promoter. Inhibition of endogenous Rab27 function in primary megakaryocytes causes severe quantitative and qualitative defects in proplatelet formation that mimic findings in gm/gm cells. Rab27b localizes to alpha and dense granules in megakaryocytes. These results establish a role for Rab27 in platelet synthesis and suggest that Rab27b in particular may coordinate proplatelet formation with granule transport, possibly by recruiting specific effector pathways.
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The melanosome: membrane dynamics in black and white
Nature Reviews Molecular Cell Biology, 2001Co-Authors: Michael S. Marks, Miguel C. SeabraAbstract:Melanosomes are morphologically and functionally unique organelles within which melanin pigments are synthesized and stored. Melanosomes share some characteristics with lysosomes, but can be distinguished from them in many ways. The biogenesis and intracellular movement of melanosomes and related organelles are disrupted in several genetic disorders in mice and humans. The recent characterization of genes defective in these diseases has reinvigorated interest in the melanosome as a model system for understanding the molecular mechanisms that underlie intracellular membrane dynamics. Melanosomes are specialized organelles within melanocytes and retinal pigment epithelial cells, where melanins, the major pigments made by mammals, are synthesized and stored. Melanosomes are one of several "lysosome-related" organelles singularly expressed in various tissues; these diverse organelles display unique morphological and functional characteristics but also share features with conventional lysosomes. Lysosome-related organelles, including melanosomes, are functionally disrupted in a group of genetic disorders in humans and mice, including Hermansky-Pudlak, Chediak-Higashi, and Griscelli syndromes. These disorders are due to defects in Protein transport, morphogenesis, or intracellular movement of lysosome-related organelles. Melanosomes serve as the best current model system in which to define the molecular basis of the disease-associated defects. Among the gene products deficient in Hermansky-Pudlak syndrome and related disorders are several involved in vesicular transport — including the adaptor complex AP-3, the SNARE-associated Protein palladin, and the α-subunit of Rab geranylgeranyltransferase. Testable models for how these and other less-characterized disease-associated Proteins function in melanosome biogenesis are being developed and are based on recent advances in our understanding of the endosomal origins of melanosome precursors, the role of multivesicular bodies, and the segregation of melanosomes from lysosomes. Morphological and biochemical analyses of melanosome resident Proteins in melanocytes from affected mice and individuals are beginning to refine these models. Griscelli's syndrome and related disorders in humans and mice result from defects in the intracellular movement and distribution of melanosomes and other lysosome-related organelles. Associated gene products regulate capture of melanosomes in the periphery of melanocytes, which are required for subsequent transfer of melanin to keratinocytes. Study of these gene products and the effects of their loss have provided new paradigms for the roles of a Rab Protein (Rab27a), a Rab effector Protein (melanophilin), and an unconventional myosin (myosin Va) in mediating actin-dependent organelle movement. Further study of melanosome biology in normal and diseased cells is likely to provide us with new paradigms to explain how conserved mechanisms are manipulated to effect the generation of structurally and functionally unique organelles and how intracellular organelle movement and positioning is regulated.
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Rab ESCORT Protein-1 IS A MULTIFUNCTIONAL Protein THAT ACCOMPANIES NEWLY PRENYLATED Rab ProteinS TO THEIR TARGET MEMBRANES
The EMBO journal, 1994Co-Authors: Kirill Alexandrov, Miguel C. Seabra, Hisanori Horiuchi, Olivia Steele-mortimer, Marino ZerialAbstract:Rab Proteins comprise a family of small GTPases that serve a regulatory role in vesicular membrane traffic. Geranylgeranylation of these Proteins on C-terminal cysteine motifs is crucial for their membrane association and function. This post-translational modification is catalysed by Rab geranylgeranyl transferase (Rab-GGTase), a multisubunit enzyme consisting of a catalytic heterodimer and an accessory component, named Rab escort Protein (REP)-1. Previous in vitro studies have suggested that REP-1 presents newly synthesized Rab Proteins to the catalytic component of the enzyme, and forms a stable complex with the prenylated Proteins following the transfer reaction. According to this model, a cellular factor would be required to dissociate the Rab Protein from REP-1 and to allow it to recycle in the prenylation reaction. RabGDP dissociation inhibitor (RabGDI) was considered an ideal candidate for this role, given its established function in mediating membrane association of prenylated Rab Proteins. Here we demonstrate that dissociation from REP-1 and binding of Rab Proteins to the membrane do not require RabGDI or other cytosolic factors. The mechanism of REP-1-mediated membrane association of Rab5 appears to be very similar to that mediated by RabGDI. Furthermore, REP-1 and RabGDI share several other functional properties, the ability to inhibit the release of GDP and to remove Rab Proteins from membranes; however, RabGDI cannot assist in the prenylation reaction. These data suggest that REP-1 is per se sufficient to chaperone newly prenylated Rab Proteins to their target membranes.
Suzanne R Pfeffer - One of the best experts on this subject based on the ideXlab platform.
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LRRK2 and Rab GTPases
Biochemical Society Transactions, 2018Co-Authors: Suzanne R PfefferAbstract:Leucine-rich repeat kinase 2 (LRRK2) is mutated in familial Parkinson9s disease, and pathogenic mutations activate the kinase activity. A tour de force screen by Mann and Alessi and co-workers identified a subset of Rab GTPases as bona fide LRRK2 substrates. Rab GTPases are master regulators of membrane trafficking and this short review will summarize what we know about the connection between LRRK2 and this family of regulatory Proteins. While, in most cases, Rab GTPase phosphorylation is predicted to interfere with Rab Protein function, the discovery of Proteins that show preferential binding to phosphorylated Rabs suggests that more complex interactions may also contribute to mutant LRRK2-mediated pathology.
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systematic proteomic analysis of lrrk2 mediated Rab gtpase phosphorylation establishes a connection to ciliogenesis
eLife, 2017Co-Authors: Martin Steger, Francesca Tonelli, Federico Diez, Suzanne R Pfeffer, Terina N Martinez, Herschel S Dhekne, Raja S Nirujogi, Ozge Karayel, Esben Lorentzen, Dario R AlessiAbstract:We previously reported that Parkinson’s disease (PD) kinase LRRK2 phosphorylates a subset of Rab GTPases on a conserved residue in their switch-II domains (Steger et al., 2016) (PMID: 26824392). Here, we systematically analyzed the Rab Protein family and found 14 of them (Rab3A/B/C/D, Rab5A/B/C, Rab8A/B, Rab10, Rab12, Rab29, Rab35 and Rab43) to be specifically phosphorylated by LRRK2, with evidence for endogenous phosphorylation for ten of them (Rab3A/B/C/D, Rab8A/B, Rab10, Rab12, Rab35 and Rab43). Affinity enrichment mass spectrometry revealed that the primary ciliogenesis regulator, RILPL1 specifically interacts with the LRRK2-phosphorylated forms of Rab8A and Rab10, whereas RILPL2 binds to phosphorylated Rab8A, Rab10, and Rab12. Induction of primary cilia formation by serum starvation led to a two-fold reduction in ciliogenesis in fibroblasts derived from pathogenic LRRK2-R1441G knock-in mice. These results implicate LRRK2 in primary ciliogenesis and suggest that Rab-mediated Protein transport and/or signaling defects at cilia may contribute to LRRK2-dependent pathologies.
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Regulation of Receptor Trafficking by ras-Like GTPases
Alzheimer’s Disease: Lessons from Cell Biology, 1995Co-Authors: Suzanne R PfefferAbstract:The trafficking of Proteins, such as the amyloid precursor, between the Golgi complex, the cell surface, and the endocytic compartment is regulated by a family of ras-like GTPases termed “Rab” Proteins. We are trying to understand the mechanism by which Rab Proteins regulate receptor trafficking. We study Rab9 and its role in facilitating the transport of Proteins between late endosomes and the trans Golgi network. Stable expression of a dominant negative mutant form of Rab9 Protein, Rab9 S21N, strongly inhibited the transport of mannose 6-phosphate receptors from late endosomes to the trans Golgi network in cultured cells. When this pathway is blocked, other aspects of endocytosis and receptor trafficking proceed normally. In addition, transport vesicle formation appears to require incorporation of a Rab Protein in its GTP-bound conformation, since receptors were not trapped in nonfunctional transport vesicles in cells expressing Rab9 S21N Protein. This is a surprise, since Rab Proteins are believed to facilitate transport vesicle targeting and/or fusion. Our findings suggest that transport vesicle formation is regulated by the availability of transport factors assembled in some fashion on the surface of newly forming transport vesicles.
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Rab gdi a solubilizing and recycling factor for Rab9 Protein
Molecular Biology of the Cell, 1993Co-Authors: Thierry Soldati, M A Riederer, Suzanne R PfefferAbstract:Rab Proteins are thought to function in the processes by which transport vesicles identify and/or fuse with their respective target membranes. The bulk of these Proteins are membrane associated, but a measuRable fraction can be found in the cytosol. The cytosolic forms of Rab3A, Rab11, and Sec4 occur as equimolar complexes with a class of Proteins termed "GDIs," or "GDP dissociation inhibitors." We show here that the cytosolic form of Rab9, a Protein required for transport between late endosomes and the trans Golgi network, also occurs as a complex with a GDI-like Protein, with an apparent mass of approximately 80 kD. Complex formation could be reconstituted in vitro using recombinant Rab9 Protein, cytosol, ATP, and geranylgeranyl diphosphate, and was shown to require an intact Rab9 carboxy terminus, as well as Rab9 geranylgeranylation. Monoprenylation was sufficient for complex formation because a mutant Rab9 Protein bearing the carboxy terminal sequence, CLLL, was prenylated in vitro by geranylgeranyl transferase I and was efficiently incorporated into 80-kD complexes. Purified, prenylated Rab9 could also assemble into 80-kD complexes by addition of purified, Rab3A GDI. Finally, Rab3A-GDI had the capacity to solubilize Rab9GDP, but not Rab9GTP, from cytoplasmic membranes. These findings support the proposal that GDI Proteins serve to recycle Rab Proteins from their target membranes after completion of a Rab Protein-mediated, catalytic cycle. Thus GDI Proteins have the potential to regulate the availability of specific intracellular transport factors.
Bruno Goud - One of the best experts on this subject based on the ideXlab platform.
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an arf6 Rab35 gtpase cascade for endocytic recycling and successful cytokinesis
Current Biology, 2012Co-Authors: Laurent Chesneau, Mitsunori Fukuda, Bruno Goud, Daphne Dambournet, Mickael Machicoane, Ilektra Kouranti, Arnaud EchardAbstract:Summary Cytokinesis bridge instability leads to binucleated cells that can promote tumorigenesis in vivo [1]. Membrane trafficking is crucial for animal cell cytokinesis [2–8], and several endocytic pathways regulated by distinct GTPases (Rab11, Rab21, Rab35, ARF6, RalA/B) [9–16] contribute to the postfurrowing steps of cytokinesis. However, little is known about how these pathways are coordinated for successful cytokinesis. The Rab35 GTPase controls a fast endocytic recycling pathway and must be activated for SEPTIN cytoskeleton localization at the intercellular bridge, and thus for completion of cytokinesis [12]. Here, we report that the ARF6 GTPase [17, 18] negatively regulates Rab35 activation and hence the Rab35 pathway. Human cells expressing a constitutively activated, GTP-bound ARF6 mutant display identical endocytic recycling and cytokinesis defects as those observed upon overexpression of the inactivated, GDP-bound Rab35 mutant. As a molecular mechanism, we identified the Rab35 GAP EPI64B as an effector of ARF6 in negatively regulating Rab35 activation. Unexpectedly, this regulation takes place at clathrin-coated pits, and activated ARF6 reduces Rab35 loading into the endocytic pathway. Thus, an effector of an ARF Protein is a GAP for a downstream Rab Protein, and we propose that this hierarchical ARF/Rab GTPase cascade controls the proper activation of a common endocytic pathway essential for cytokinesis.
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Rab6 and the secretory pathway affect oocyte polarity in Drosophila.
Development (Cambridge England), 2007Co-Authors: Jens Januschke, Bruno Goud, Emmanuelle Nicolas, Julien Compagnon, Etienne Formstecher, Antoine GuichetAbstract:The Drosophila oocyte is a highly polarized cell. Secretion occurs towards restricted neighboring cells and asymmetric transport controls the localization of several mRNAs to distinct cortical compartments. Here, we describe a role for the Drosophila ortholog of the Rab6 GTPase, DRab6, in establishing cell polarity during oogenesis. We found that DRab6 localizes to Golgi and Golgi-derived membranes and interacts with BicD. We also provide evidence that DRab6 and BicD function together to ensure the correct delivery of secretory pathway components, such as the TGFalpha homolog Gurken, to the plasma membrane. Moreover, in the absence of DRab6, osk mRNA localization and the organization of microtubule plus-ends at the posterior of the oocyte were both severely affected. Our results point to a possible connection between Rab Protein-mediated secretion, organization of the cytoskeleton and mRNA transport.
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Selection and application of recombinant antibodies as sensors of Rab Protein conformation.
Methods in enzymology, 2005Co-Authors: Clément Nizak, Sandrine Moutel, Bruno Goud, Franck PerezAbstract:Abstract The existence of a conformational switch of Rabs and other small GTPases involved in intracellular transport regulation has been known for many years. This switch is superimposed on the membrane association/dissociation cycle for most of these GTPases. While these processes are key features of the dynamics of intracellular transport events, surprisingly very few previous studies have focused on the dynamics of the GDP/GTP cycle of Rab Proteins in time and space. The main reason for this is the lack of tools available to dynamically probe for Rab GTPases conformation switches and membrane association/dissociation, in particular in vivo . We recently reported the in vitro selection of conformation‐specific recombinant antibodies specific to the GTP‐bound conformation of Rab6 Proteins. These antibodies were obtained in vitro by phage display, a rather simple, rapid, and cheap technique. We additionally showed that these conformation‐specific antibodies can be expressed in living cells to follow endogenous Rab6 in its activated conformation in vivo . The same strategy could be used to study other conformation switching mechanisms and, in general, to study the switching between states that antibodies can distinguish (e.g., phosphorylation, ubiquitination).
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Multiple aspects of Rab Protein action in the secretory pathway: focus on Rab3 and Rab6.
Biochimie, 2000Co-Authors: François Darchen, Bruno GoudAbstract:Rab Proteins form the largest branch of the Ras superfamily of GTPases. They are localized to the cytoplasmic face of organelles and vesicles involved in the biosynthetic/secretory and endocytic pathways in eukaryotic cells. It is now well established that Rab Proteins play an essential role in the processes that underlie the targeting and fusion of transport vesicles with their appropriate acceptor membranes. They perform this task through interactions with a wide variety of effector molecules. In this review, we illustrate recent advances in the field of Rab GTPases, taking as examples two Proteins involved in the biosynthetic pathway, Rab3 and Rab6.
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interaction of a golgi associated kinesin like Protein with Rab6
Science, 1998Co-Authors: Arnaud Echard, Florence Jollivet, Olivier Martinez, Jeanjacques Lacapere, Annie Rousselet, Isabelle Janoueixlerosey, Bruno GoudAbstract:Rab guanosine triphosphatases regulate vesicular transport and membrane traffic within eukaryotic cells. Here, a kinesin-like Protein that interacts with guanosine triphosphate (GTP)-bound forms of Rab6 was identified. This Protein, termed Rabkinesin-6, was localized to the Golgi apparatus and shown to play a role in the dynamics of this organelle. The carboxyl-terminal domain of Rabkinesin-6, which contains the Rab6-interacting domain, inhibited the effects of Rab6-GTP on intracellular transport. Thus, a molecular motor is a potential effector of a Rab Protein, and coordinated action between members of these two families of Proteins could control membrane dynamics and directional vesicular traffic.
