Identification of Endosomal Epidermal Growth Factor Receptor Signaling Targets by Functional Organelle Proteomics

Stasyk, Taras; Schiefermeier, Natalia; Skvortsov, Sergej; Zwierzina, Heinz; Peränen, Johan; Bonn, Guenther K.; Huber, Lukas A.

doi:10.1074/mcp.m600463-mcp200

Cited by 53 publications

(44 citation statements)

References 60 publications

(89 reference statements)

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“…In this study we extended earlier observations on endosomal events after EGF treatment (6,10,15,66). Of particular note was the finding that EGF induced the accumulation of V 1 components of the V-ATPase in endosomal DRMs/rafts, whereas the V 0 components showed little or no change in this fraction.…”

Section: Discussionsupporting

confidence: 72%

Epidermal Growth Factor-induced Vacuolar (H+)-ATPase Assembly

Parmar

Roux

et al. 2012

Journal of Biological Chemistry

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Background: EGF-induced activation of mTORC1 in hepatocytes is essential for cell proliferation. Results: Blocking EGF-induced vacuolar acidification reduced intracellular essential amino acid levels and inhibited mTORC1 signaling without affecting Akt/Erk activation. Conclusion: EGF-induced mTORC1 signaling depends on sustaining intracellular amino acid levels. Significance: Our results support a role for vacuolar acidification in growth factor signaling.

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Section: Discussionsupporting

confidence: 72%

Epidermal Growth Factor-induced Vacuolar (H+)-ATPase Assembly

Parmar

Roux

et al. 2012

Journal of Biological Chemistry

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“…The EEA1-positive early endosomes were isolated as previously described [28,29]. In brief, the cells were washed with icecold PBS twice, rinsed with osmotic buffer (10 mM Tris/HCl, pH 7.4) for 1 min, and treated with homogenization buffer (10 mM Tris/HCl, 1 mM EGTA, 0.5 mM EDTA, 3 mM Imidazole, 250 mM sucrose, Complete protease inhibitors, pH 7.4).…”

Section: Immunoisolation Of Early Endosomesmentioning

confidence: 99%

“…By labeling EEA1 and caveolin-1 using different sizes of gold nanoparticles, we found the distribution of caveolin-1 on the membrane of EEA1-positive early endosomes (Supplementary information, Figure S6). (3) We immunoisolated the EEA1-positive early endosomes using a subcellular fractionation protocol [28,29]. The immunoisolated early endosomes were then analyzed by western blotting with antibodies against EEA1, caveolin-1, TGF-β receptor, Golgi protein p115 and mitochondrial protein Tim23.…”

Section: Intracellular Distribution Of Tgf-β Receptors In Caveolin-1-mentioning

confidence: 99%

Internalization of the TGF-β type I receptor into caveolin-1 and EEA1 double-positive early endosomes

Yan

et al. 2015

Cell Res

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Endocytosis and intracellular sorting of transforming growth factor-β (TGF-β) receptors play an important regulatory role in TGF-β signaling. Two major endocytic pathways, clathrin-and caveolae-mediated endocytosis, have been reported to independently mediate the internalization of TGF-β receptors. In this study, we demonstrate that the clathrin-and caveolae-mediated endocytic pathways can converge during TGF-β receptor endocytic trafficking. By tracking the intracellular dynamics of fluorescently-labeled TGF-β type I receptor (TβRI), we found that after mediating TβRI internalization, certain clathrin-coated vesicles and caveolar vesicles are fused underneath the plasma membrane, forming a novel type of caveolin-1 and clathrin double-positive vesicles. Under the regulation of Rab5, the fused vesicles are targeted to early endosomes and thus deliver the internalized TβRI to the caveolin-1 and EEA1 double-positive early endosomes (caveolin-1-positive early endosomes). We further showed that the caveolin-1-positive early endosomes are positive for Smad3/SARA, Rab11 and Smad7/Smurf2, and may act as a multifunctional device for TGF-β signaling and TGF-β receptor recycling and degradation. Therefore, these findings uncover a novel scenario of endocytosis, the direct fusion of clathrin-coated and caveolae vesicles during TGF-β receptor endocytic trafficking, which leads to the formation of the multifunctional sorting device, caveolin-1-positive early endosomes, for TGF-β receptors.

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“…2 and J. Mankouri, S. Griffin & M. Harris, unpublished) and that of others (Tang et al, 2007) suggest that HCV NS5A can colocalize with markers of early endosomes such as Rab5 and EEA1. It is tempting to speculate that the ability of HCV NS5A to localize to such compartments may be related to its ability to inhibit Ras-Erk signalling, particularly in the light of studies showing that EGFR signalling events require receptor endocytosis into endosomal compartments (Stasyk et al, 2007;Vieira et al, 1996).…”

Section: Discussionmentioning

confidence: 99%

A comparative cell biological analysis reveals only limited functional homology between the NS5A proteins of hepatitis C virus and GB virus B

Mankouri

Milward

Pryde

et al. 2008

Journal of General Virology

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GB virus B (GBV-B) is the closest relative to hepatitis C virus (HCV) with which it shares a common genome organization, however, unlike HCV in humans, it generally causes an acute resolving hepatitis in New World monkeys. It is important to understand the factors regulating the different disease profiles of the two viruses and in this regard, as well as playing a key role in viral RNA replication, the HCV NS5A non-structural protein modulates a variety of host-cell signalling pathways. We have shown previously that HCV NS5A, expressed either alone, or in the context of the complete polyprotein, inhibits the Ras-extracellular-signal-regulated kinase (Erk) pathway and activates the phosphoinositide 3-kinase (PI3K) pathway. In this report, we investigate whether these functions are shared by GBV-B NS5A. Immunofluorescence analysis revealed that a C-terminally FLAG-tagged GBV-B NS5A exhibited a punctate cytoplasmic distribution. However, unlike HCV NS5A, the GBV-B protein did not partially co-localize with early endosomes. Utilizing a transient luciferase reporter system, we observed that GBV-B NS5A failed to inhibit Ras-Erk signalling, however GBV-B NS5A expression did result in the elevation of b-catenin-dependent transcription via activation of the PI3K pathway. These effects of GBV-B and HCV NS5A on the PI3K and Ras-Erk pathways were confirmed in cells harbouring subgenomic replicons derived from the two viruses. Based on these data we speculate that the differential effects of the two NS5A proteins on cellular signalling pathways may contribute to the differences in the natural history of the two viruses. INTRODUCTIONGB virus B (GBV-B) is the closest phylogenetic relative of hepatitis C virus (HCV) (Muerhoff et al., 1995;Ohba et al., 1996); it shares up to 28 % overall amino acid identity with HCV, rising to 50 % conservation in regions such as the NS3 coding sequence. GBV-B has been proposed as a model for HCV infection and has been used to characterize candidate antivirals as well as for pathogenesis studies (Beames et al., 2001;Bright et al., 2004). It shares the hepatotropism of HCV. Although GBV-B has been reported to lead to persistent infections in some cases in tamarins (Saguinus species) (Martin et al., 2003;Nam et al., 2004), unlike the human virus, GBV-B primarily causes an acute, resolving hepatitis in tamarins and other New World monkeys such as common marmosets (Callithrix jacchus) (Bukh et al., 2001;Jacob et al., 2004;Lanford et al., 2003). In contrast, HCV is a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma, affecting an estimated 3 % of the world's population.Both GBV-B and HCV are members of the genus Hepacivirus of the family Flaviviridae and have a positivesense RNA genome that is translated in a cap-independent fashion to generate a polyprotein, cleaved by host and viral proteases to produce 10 mature viral proteins. There are three structural proteins at the N terminus: core, E1 and E2, followed by a small cation channel, p7 (p13 in GBV-B). The C-terminal t...

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Identification of Endosomal Epidermal Growth Factor Receptor Signaling Targets by Functional Organelle Proteomics

Cited by 53 publications

References 60 publications

Epidermal Growth Factor-induced Vacuolar (H+)-ATPase Assembly

Epidermal Growth Factor-induced Vacuolar (H+)-ATPase Assembly

Internalization of the TGF-β type I receptor into caveolin-1 and EEA1 double-positive early endosomes

A comparative cell biological analysis reveals only limited functional homology between the NS5A proteins of hepatitis C virus and GB virus B

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