| 1. |
- Forsmark, Annabelle, 1973, et al.
(författare)
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Quantitative Proteomics of Yeast Post-Golgi Vesicles Reveals a Discriminating Role for Sro7p in Protein Secretion
- 2011
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Ingår i: Traffic. - : John Wiley & Sons. - 1398-9219 .- 1600-0854. ; 12:6, s. 740-753
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Tidskriftsartikel (refereegranskat)abstract
- We here report the first comparative proteomics of purified yeast post-Golgi vesicles (PGVs). Vesicle samples isolated from PGV-accumulating sec6-4 mutants were treated with isobaric tags (iTRAQ) for subsequent quantitative tandem mass spectrometric analysis of protein content. After background subtraction, a total of 66 vesicle-associated proteins were identified, including known or assumed vesicle residents as well as a fraction not previously known to be PGV associated. Vesicles isolated from cells lacking the polarity protein Sro7p contained essentially the same catalogue of proteins but showed a reduced content of a subset of cargo proteins, in agreement with a previously shown selective role for Sro7p in cargo sorting.
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| 2. |
- Nilsson, Lars, et al.
(författare)
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Caenorhabditis elegans Numb Inhibits Endocytic Recycling by Binding TAT-1 Aminophospholipid Translocase
- 2011
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Ingår i: Traffic. - Malden : Wiley-Blackwell. - 1398-9219 .- 1600-0854. ; 12:12, s. 1839-1849
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Tidskriftsartikel (refereegranskat)abstract
- Numb regulates endocytosis in many metazoans, but the mechanism by which it functions is not completely understood. Here we report that the Caenorhabditis ele-gans Numb ortholog, NUM-1A, a regulator of endocytic recycling, binds the C isoform of transbilayer amphipath transporter-1 (TAT-1), a P4 family adenosine triphosphatase and putative aminophospholipid translocase that is required for proper endocytic trafficking. We demonstrate that TAT-1 is differentially spliced during development and that TAT-1C-specific splicing occurs in the intestine where NUM-1A is known to function. NUM-1A and TAT-1C colocalize in vivo. We have mapped the binding site to an NXXF motif in TAT-1C. This motif is not required for TAT-1C function but is required for NUM-1A's ability to inhibit recycling. We demonstrate that num-1A and tat-1 defects are both suppressed by the loss of the activity of PSSY-1, a phosphatidylserine (PS) synthase. PS is mislocalized in intestinal cells with defects in tat-1 or num-1A function. We propose that NUM-1A inhibits recycling by inhibiting TAT-1C's ability to translocate PS across the membranes of recycling endosomes.
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| 3. |
- Pattu, Varsha, et al.
(författare)
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Syntaxin7 is required for lytic granule release from cytotoxic T lymphocytes.
- 2011
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Ingår i: Traffic. - : Wiley. - 1398-9219 .- 1600-0854. ; 12:7
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Tidskriftsartikel (refereegranskat)abstract
- SNARE proteins are essential fusion mediators for many intracellular trafficking events. Here, we investigate the role of Syntaxin7 (Stx7) in the release of lytic granules from cytotoxic T lymphocytes (CTLs). We show that Stx7 is expressed in CTLs and is preferentially localized to the region of lytic granule release, the immunological synapse (IS). Interference of Stx7 function by expression of a dominant-negative Stx7 construct or by small interfering RNA leads to a dramatic reduction of CTL-mediated killing of target cells. Real-time visualization of individual lytic granules at the IS by evanescent wave microscopy reveals that lytic granules in Stx7-deprived CTLs not only fail to fuse with the plasma membrane but even fail to accumulate at the IS. Surprisingly, the accumulation defect is not caused by an overall reduction in lytic granule number, but by a defect in the trafficking of T cell receptors (TCRs) through endosomes. Subsequent high-resolution nanoscopy shows that Stx7 colocalizes with Rab7 on late endosomes. We conclude from these data that the accumulation of recycling TCRs at the IS is a SNARE-dependent process and that Stx7-mediated processing of recycling TCRs through endosomes is a prerequisite for the cytolytic function of CTLs.
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| 4. |
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| 5. |
- Dudenhöffer-Pfeifer, Monika, et al.
(författare)
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Different Munc13 isoforms function as priming factors in lytic granule release from murine cytotoxic T lymphocytes.
- 2013
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Ingår i: Traffic. - : Wiley. - 1398-9219 .- 1600-0854. ; 14:7
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Tidskriftsartikel (refereegranskat)abstract
- In order to fuse lytic granules (LGs) with the plasma membrane at the immunological synapse, cytotoxic T lymphocytes (CTLs) have to render these LGs fusion-competent through the priming process. In secretory tissues such as brain and neuroendocrine glands, this process is mediated by members of the Munc13 protein family. In human CTLs, mutations in the Munc13-4 gene cause a severe loss in killing efficiency, resulting in familial hemophagocytic lymphohistiocytosis type 3, suggesting a similar role of other Munc13 isoforms in the immune system. Here, we investigate the contribution of different Munc13 isoforms to the priming process of murine CTLs at both the mRNA and protein level. We demonstrate that Munc13-1 and Munc13-4 are the only Munc13 isoforms present in mouse CTLs. Both isoforms rescue the drastical secretion defect of CTLs derived from Munc13-4-deficient Jinx mice. Mobility studies using total internal reflection fluorescence microscopy indicate that Munc13-4 and Munc13-1 are responsible for the priming process of LGs. Furthermore, the domains of the Munc13 protein, which is responsible for functional fusion, could be identified. We conclude from these data that both isoforms of the Munc13 family, Munc13-1 and Munc13-4, are functionally redundant in murine CTLs.
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| 6. |
- Frénal, Karine, et al.
(författare)
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Global analysis of apicomplexan protein S-acyl transferases reveals an enzyme essential for invasion
- 2013
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Ingår i: Traffic. - : John Wiley & Sons. - 1398-9219 .- 1600-0854. ; 14:8, s. 895-911
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Tidskriftsartikel (refereegranskat)abstract
- The advent of techniques to study palmitoylation on a whole proteome scale has revealed that it is an important reversible modification that plays a role in regulating multiple biological processes. Palmitoylation can control the affinity of a protein for lipid membranes, which allows it to impact protein trafficking, stability, folding, signalling and interactions. The publication of the palmitome of the schizont stage of Plasmodium falciparum implicated a role for palmitoylation in host cell invasion, protein export and organelle biogenesis. However, nothing is known so far about the repertoire of protein S-acyl transferases (PATs) that catalyse this modification in Apicomplexa. We undertook a comprehensive analysis of the repertoire of Asp-His-His-Cys cysteine-rich domain (DHHC-CRD) PAT family in Toxoplasma gondii and Plasmodium berghei by assessing their localization and essentiality. Unlike functional redundancies reported in other eukaryotes, some apicomplexan-specific DHHCs are essential for parasite growth, and several are targeted to organelles unique to this phylum. Of particular interest is DHHC7, which localizes to rhoptry organelles in all parasites tested, including the major human pathogen P. falciparum. TgDHHC7 interferes with the localization of the rhoptry palmitoylated protein TgARO and affects the apical positioning of the rhoptry organelles. This PAT has a major impact on T. gondii host cell invasion, but not on the parasite's ability to egress.
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| 7. |
- Nehru, Vishal, et al.
(författare)
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RhoD Binds the Rab5 Effector Rabankyrin-5 and has a Role in Trafficking of the Platelet-derived Growth Factor Receptor
- 2013
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Ingår i: Traffic. - : Wiley. - 1398-9219 .- 1600-0854. ; 14:12, s. 1242-1254
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Tidskriftsartikel (refereegranskat)abstract
- RhoD is a member of the classical Rho GTPases and it has essential roles in the regulation of actin dynamics. RhoD localizes to early endosomes and recycling endosomes, which indicates its important role in the regulation of endosome trafficking. Here, we show that RhoD binds to the Rab5 effector Rabankyrin-5, and RhoD and Rabankyrin-5 colocalize to Rab5-positive endosomes, which suggests a role for Rabankyrin-5 in the coordination of RhoD and Rab5 in endosomal trafficking. Interestingly, depletion of RhoD using siRNA techniques interfered with the internalization of the PDGF receptor and the subsequent activation of the downstream signaling cascades. Our data suggest that RhoD and Rabankyrin-5 have important roles in coordinating RhoD and Rab activities during internalization and trafficking of activated tyrosine kinase receptors.
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| 8. |
- Sadowski, Lukasz, et al.
(författare)
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Dynamin Inhibitors Impair Endocytosis and Mitogenic Signaling of PDGF
- 2013
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Ingår i: Traffic. - : John Wiley & Sons. - 1398-9219 .- 1600-0854. ; 14:6, s. 725-736
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Tidskriftsartikel (refereegranskat)abstract
- Platelet-derived growth factor (PDGF) isoforms regulate cell proliferation, migration and differentiation both in embryonic development and adult tissue remodeling. At the cellular level, growth-factor signaling is often modulated by endocytosis. Despite important functions of PDGF, its endocytosis remains poorly studied, mainly for lack of tools to track internalized ligand by microscopy. Here, we developed such a tool and quantitatively analyzed internalization and endosomal trafficking of PDGF-BB in human fibroblasts. We further show that PDGF can be internalized in the presence of dynamin inhibitors, arguing that both dynamin-dependent and dynamin-independent pathways can mediate PDGF uptake. Although these routes operate with somewhat different kinetics, they both ultimately lead to lysosomal degradation of PDGF. Although acute inhibition of dynamin activity only moderately affects PDGF endocytosis, it specifically decreases downstream signaling of PDGF via signal transducer and activator of transcription 3 (STAT3). This correlates with reduced expression of MYC and impaired cell entry into S-phase, indicating that dynamin activity is required for PDGF-induced mitogenesis. Our data support a general view that the components governing endocytic trafficking may selectively regulate certain signaling effectors activated by a growth factor.
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| 9. |
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