172,807 research outputs found
Structure, regulation and cellular functions of Rab geranylgeranyl transferase and its cellular partner Rab Escort Protein
No full textRab geranylgeranyl transferase is an enzyme responsible for double geranylgeranylation of Rab proteins in all eukaryotic cells. In the present article we would like to focus on new findings concerning the holoenzyme structure and mechanism of catalytic activity, its mode of regulation and consequences of RGGT deficiency in different eucaryotic model organisms and patients
Specific Rab GTPase-activating proteins define the Shiga toxin and epidermal growth factor uptake pathways
No full textRab family guanosine triphosphatases (GTPases) together with their regulators define specific pathways of membrane traffic within eukaryotic cells. In this study, we have investigated which Rab GTPase-activating proteins (GAPs) can interfere with the trafficking of Shiga toxin from the cell surface to the Golgi apparatus and studied transport of the epidermal growth factor (EGF) from the cell surface to endosomes. This screen identifies 6 (EVI5, RN-tre/USP6NL, TBC1D10A–C, and TBC1D17) of 39 predicted human Rab GAPs as specific regulators of Shiga toxin but not EGF uptake. We show that Rab43 is the target of RN-tre and is required for Shiga toxin uptake. In contrast, RabGAP-5, a Rab5 GAP, was unique among the GAPs tested and reduced the uptake of EGF but not Shiga toxin. These results suggest that Shiga toxin trafficking to the Golgi is a multistep process controlled by several Rab GAPs and their target Rabs and that this process is discrete from ligand-induced EGF receptor trafficking
RAB-5 and RAB-7 GTPases promote DAF-16 FOXO localization to endosomes.
No full text(A-F) Representative confocal images of the intestinal cells of animals expressing DAF-16a::GFP (zIs356) (green) and either RFP::RAB-5 (pwIs480) (A-C) or mCherry::RAB-7 (pwIs429) (magenta) (D- F). Arrowheads mark examples of vesicles positive for both DAF-16a::GFP and either RFP::RAB-5 or mCherry::RAB-7. Arrows mark examples of DAF-16a::GFP vesicles that are not positive for either RFP::RAB-5 or mCherry::RAB-7. (G and H) Bar graphs displaying the mean (and SEM) of the percent wild-type and tbc-2(tm2241) animals with DAF-16a::GFP (zIs356) vesicles fed bacteria expressing control empty vector (ev) RNAi (black bars) compared to animals fed rab-5(RNAi) or rab-7(RNAi) (white bars) from three independent experiments. Raw data is available in S1 Data. Fisher’s exact test (graphpad.com) was used to determine the statistical difference between conditions. n, total number of animals. * P.05, *** P.001, **** P.0001.</p
RAB-28 undergoes IFT.
No full text(A, B) Representative images of phasmid cilia (left) from N2 wild type and che-11(e1810) worms expressing GFP::RAB-28, together with corresponding kymographs and kymograph schematics derived from time-lapse imaging. Distribution plot (and mean values) in B shows kymography-determined anterograde and retrograde GFP::RAB-28 velocities from wild type worms. MS; middle segment, DS; distal segment, PC; periciliary membrane compartment, DD; distal dendrite. Scale bars; 3 μm (phasmid image; and horizontal bar on kymographs); 3 seconds (vertical bar on kymographs). (C) Fluorescence recovery after photobleaching (FRAP) plots for GFP::RAB-28 in the phasmid neurons of N2 and che-11(e1810) mutant worms. GFP::RAB-28 also undergoes free diffusion in wild-type (N2) and che-11 IFT mutant animals. Ciliary GFP signals bleached at time 0. Intensity measurements normalised to pre-bleach levels. Curves derived from 3 separate FRAP experiments. Error bars; SEM. Images taken from a representative FRAP experiment in N2 wild type worms. s; seconds. Scale bar; 2 μm.</p
Construction of a Plasmodium falciparum Rab-interactome identifies CK1 and PKA as Rab-effector kinases in malaria parasites
No full textBackground information
The pathology causing stages of the human malaria parasite Plasmodium falciparum reside within red blood cells that are devoid of any regulated transport system. The parasite, therefore, is entirely responsible for mediating vesicular transport within itself and in the infected erythrocyte cytoplasm, and it does so in part via its family of 11 Rab GTPases. Putative functions have been ascribed to Plasmodium Rabs due to their homology with Rabs of yeast, particularly with Saccharomyces that has an equivalent number of rab/ypt genes and where analyses of Ypt function is well characterized.
Results
Rabs are important regulators of vesicular traffic due to their capacity to recruit specific effectors. In order to identify P. falciparum Rab (PfRab) effectors, we first built a Ypt-interactome by exploiting genetic and physical binding data available at the Saccharomyces genome database (SGD). We then constructed a PfRab-interactome using putative parasite Rab-effectors identified by homology to Ypt-effectors. We demonstrate its potential by wet-bench testing three predictions; that casein kinase-1 (PfCK1) is a specific Rab5B interacting protein and that the catalytic subunit of cAMP-dependent protein kinase A (PfPKA-C) is a PfRab5A and PfRab7 effector.
Conclusions
The establishment of a shared set of physical Ypt/PfRab-effector proteins sheds light on a core set Plasmodium Rab-interactants shared with yeast. The PfRab-interactome should benefit vesicular trafficking studies in malaria parasites. The recruitment of PfCK1 to PfRab5B+ and PfPKA-C to PfRab5A+ and PfRab7+ vesicles, respectively, suggests that PfRab-recruited kinases potentially play a role in early and late endosome function in malaria parasites
RAB-35 is localized to extending pseudopods and further enriched on nascent phagosomes.
No full textAll GFP reporters are expressed in engulfing cells under the control of Pced-1. (A) Diagram illustrating the features that help us visualize ventral enclosure and apoptotic cell clearance. The start of ventral enclosure is defined as the moment the two ventral hypodermal cells (ABpraapppp and ABplaapppp) start extending to the ventral midline. Both the position of cell corpses C1, C2, and C3 (brown dots) as well as the identity of their engulfing cells are shown. (B) Time-lapse recording of GFP::RAB-35 during the engulfment and degradation of cell corpse C3 in a wild-type embryo. “0 min”: the moment a nascent phagosome is just formed. Arrowheads mark the extending pseudopods. One arrow marks the nascent phagosome. (C) Graph showing the relative GFP::RAB-35 signal intensity over time on the surface of pseudopods and the phagosome shown in B. The GFP signal intensity was measured on the phagosomal surface and in the surrounding cytoplasm every 2 minutes, starting from the “-4 min” time point. The phagosomal / cytoplasmic signal ratio over time was presented. Data is normalized relative to the signal ratio at the “-4 min” time point. (D) Bar graph presenting the mean numbers and sd (error bars) of cell corpses scored in 1.5-fold stage wild-type or rab-35(b1013) mutant embryos, in the presence or absence of transgenes overexpressing GFP::RAB-35(S24N) or GFP::RAB-35(Q69L). For each data point, at least 15 animals were scored. Brackets above the bars indicate the samples that are compared by the Student t-test. p-values are summarized as such: *, 0.001 < p < 0.05; **, 0.00001 < p <0.001; ***, p <0.00001; ns, no significant difference. (E) Time-lapse images exhibiting the localization of GFP::RAB-35(S24N) and GFP::RAB-35(Q69L) during the engulfment and degradation of C3. “0 min” is the moment a nascent phagosome is just formed. Arrowheads mark extending pseudopods. A white arrow marks the nascent phagosome. Regions with enriched GFP::RAB-35(Q69L) signal on the phagosomal membrane are marked by yellow arrows.</p
The Great Tibetan Translator: Life and Works of rNgog Blo ldan shes rab (1059–1109)
No full textSecond only to the famous Rin chen bzang po (958–1055) in receiving the title of a “Great Translator” (lo chen) during the period of the “Later Propagation” (phyi dar) of Buddhism in Tibet, rNgog lo tsā ba Blo ldan shes rab (or rNgog lo) was one of the most influential figures in the establishment of Tibetan Buddhist scholasticism. After having devoted seventeen years of his life to the study of Sanskrit under scholars in Kashmir, India and Nepal, he became renowned for his more than fifty painstaking translations and revisions of Buddhist scriptures. Apart from being the foremost Tibetan translator of works on Buddhist logic and epistemology (Pramāṇa), rNgog lo’s activities as a commentator and teacher are regarded as fundamental for the later development of this field of learning in Tibet, and his tradition came to be well-known in Tibetan literature as the “rNgog tradition” (rngog lugs). This book presents a detailed examination of rNgog lo’s life based on the available Tibetan accounts, including his biography (rnam thar) written by Gro lung pa Blo gros ’byung gnas (fl. late 11th to 12th c.). Annotated translations of great parts from the latter work (one of the earliest surviving examples of the rnam thar genre, possibly unique regarding its complicated and elegant style) are included in the book. rNgog lo’s oeuvre as a translator and writer is dealt with in detail, making the book an important source on this hitherto little studied scholar and his tradition
RAB-6.1 and RAB-6.2 Promote Retrograde Transport in C. elegans.
No full textRetrograde transport is a critical mechanism for recycling certain membrane cargo. Following endocytosis from the plasma membrane, retrograde cargo is moved from early endosomes to Golgi followed by transport (recycling) back to the plasma membrane. The complete molecular and cellular mechanisms of retrograde transport remain unclear. The small GTPase RAB-6.2 mediates the retrograde recycling of the AMPA-type glutamate receptor (AMPAR) subunit GLR-1 in C. elegans neurons. Here we show that RAB-6.2 and a close paralog, RAB-6.1, together regulate retrograde transport in both neurons and non-neuronal tissue. Mutants for rab-6.1 or rab-6.2 fail to recycle GLR-1 receptors, resulting in GLR-1 turnover and behavioral defects indicative of diminished GLR-1 function. Loss of both rab-6.1 and rab-6.2 results in an additive effect on GLR-1 retrograde recycling, indicating that these two C. elegans Rab6 isoforms have overlapping functions. MIG-14 (Wntless) protein, which undergoes retrograde recycling, undergoes a similar degradation in intestinal epithelia in both rab-6.1 and rab-6.2 mutants, suggesting a broader role for these proteins in retrograde transport. Surprisingly, MIG-14 is localized to separate, spatially segregated endosomal compartments in rab-6.1 mutants compared to rab-6.2 mutants. Our results indicate that RAB-6.1 and RAB-6.2 have partially redundant functions in overall retrograde transport, but also have their own unique cellular- and subcellular functions
RAB-35 is enriched on phagosomal surfaces during the PtdIns(4,5)P<sub>2</sub> to PtdIns(3)P shift and contributes to the PtdIns(4,5)P<sub>2</sub> removal from nascent phagosomes.
No full text(A-C) Time-lapse images during and after the formation of a phagosome carrying C3 in wild-type embryos. “0 min” is the moment when a phagosome is just formed. Arrowheads indicate extending pseudopods. White arrows mark the nascent phagosome. (A) Reporters: Pced-1 mKate2::rab-35 and the PtdIns(4,5)P2 marker Pced-1 PH(hPLCγ)::gfp. Yellow arrows mark the moment when both the gain of mKate2::RAB-35 and the loss of PH(hPLCγ)::GFP from the phagosomal surface is observed. (B) Reporters: Pced-1 gfp::rab-35 and the PtdIns(3)P marker Pced-1 2xFYVE::mRFP. Yellow arrows mark the moment when both GFP::RAB-35 and 2xFYVE::mRFP are rapidly enriched on the phagosomal surface. (C) Reporter: Pced-1 PH(hPLCγ)::gfp. Yellow arrows mark the first time point when PtdIns(4,5)P2 is no longer observed on the phagosome surface. (D) Histograms displaying the range of time it takes for the disappearance of PtdIns(4,5)P2 from the surface of phagosomes bearing C1, C2, and C3 in embryos of various genotypes. The time interval between the formation of a nascent phagosome (“0 min”) and the first time point when the PH(hPLCγ)::GFP signal is no longer enriched on the phagosomal surface are displayed. For each genotype, at least 15 phagosomes were scored.</p
RAB-10 Promotes EHBP-1 Bridging of Filamentous Actin and Tubular Recycling Endosomes.
No full textEHBP-1 (Ehbp1) is a conserved regulator of endocytic recycling, acting as an effector of small GTPases including RAB-10 (Rab10). Here we present evidence that EHBP-1 associates with tubular endosomal phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] enriched membranes through an N-terminal C2-like (NT-C2) domain, and define residues within the NT-C2 domain that mediate membrane interaction. Furthermore, our results indicate that the EHBP-1 central calponin homology (CH) domain binds to actin microfilaments in a reaction that is stimulated by RAB-10(GTP). Loss of any aspect of this RAB-10/EHBP-1 system in the C. elegans intestinal epithelium leads to retention of basolateral recycling cargo in endosomes that have lost their normal tubular endosomal network (TEN) organization. We propose a mechanism whereby RAB-10 promotes the ability of endosome-bound EHBP-1 to also bind to the actin cytoskeleton, thereby promoting endosomal tubulation
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