scholarly journals SCAMP3 Negatively Regulates Epidermal Growth Factor Receptor Degradation and Promotes Receptor Recycling

2009 ◽  
Vol 20 (6) ◽  
pp. 1816-1832 ◽  
Author(s):  
Quyen L. Aoh ◽  
Anna M. Castle ◽  
Charles H. Hubbard ◽  
Osamu Katsumata ◽  
J. David Castle
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Growth Factor Receptor ◽  
Multivesicular Bodies ◽  
Receptor Recycling ◽  
Endosomal Sorting ◽  
Accelerated Degradation ◽  
Early Endosomes ◽  
Epidermal Growth

The epidermal growth factor receptor (EGFR) is targeted for lysosomal degradation by ubiquitin-mediated interactions with the ESCRTs (endosomal-sorting complexes required for transport) in multivesicular bodies (MVBs). We show that secretory carrier membrane protein, SCAMP3, localizes in part to early endosomes and negatively regulates EGFR degradation through processes that involve its ubiquitylation and interactions with ESCRTs. SCAMP3 is multimonoubiquitylated and is able to associate with Nedd4 HECT ubiquitin ligases and the ESCRT-I subunit Tsg101 via its PY and PSAP motifs, respectively. SCAMP3 also associates with the ESCRT-0 subunit Hrs. Depletion of SCAMP3 in HeLa cells by inhibitory RNA accelerated degradation of EGFR and EGF while inhibiting recycling. Conversely, overexpression enhanced EGFR recycling unless ubiquitylatable lysines, PY or PSAP motifs in SCAMP3 were mutated. Notably, dual depletions of SCAMP3 and ESCRT subunits suggest that SCAMP3 has a distinct function in parallel with the ESCRTs that regulates receptor degradation. This function may affect trafficking of receptors from prelysosomal compartments as SCAMP3 depletion appeared to sustain the incidence of EGFR-containing MVBs detected by immunoelectron microscopy. Together, our results suggest that SCAMP3, its modification with ubiquitin, and its interactions with ESCRTs coordinately regulate endosomal pathways and affect the efficiency of receptor down-regulation.

scholarly journals A Role for Sorting Nexin 2 in Epidermal Growth Factor Receptor Down-regulation: Evidence for Distinct Functions of Sorting Nexin 1 and 2 in Protein Trafficking

2004 ◽  
Vol 15 (5) ◽  
pp. 2143-2155 ◽  
Author(s):  
Anuradha Gullapalli ◽  
Tiana A. Garrett ◽  
May M. Paing ◽  
Courtney T. Griffin ◽  
Yonghua Yang ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Protein Trafficking ◽  
Growth Factor Receptor ◽  
Down Regulation ◽  
Sorting Nexin ◽  
Px Domain ◽  
Early Endosomes ◽  
Epidermal Growth

Sorting nexin 1 (SNX1) and SNX2, homologues of the yeast vacuolar protein-sorting (Vps)5p, contain a phospholipid-binding motif termed the phox homology (PX) domain and a carboxyl terminal coiled-coil region. A role for SNX1 in trafficking of cell surface receptors from endosomes to lysosomes has been proposed; however, the function of SNX2 remains unknown. Toward understanding the function of SNX2, we first examined the distribution of endogenous protein in HeLa cells. We show that SNX2 resides primarily in early endosomes, whereas SNX1 is found partially in early endosomes and in tubulovesicular-like structures distributed throughout the cytoplasm. We also demonstrate that SNX1 interacts with the mammalian retromer complex through its amino terminal domain, whereas SNX2 does not. Moreover, activated endogenous epidermal growth factor receptor (EGFR) colocalizes markedly with SNX2-positive endosomes, but minimally with SNX1-containing vesicles. To assess SNX2 function, we examined the effect of a PX domain-mutated SNX2 that is defective in vesicle localization on EGFR trafficking. Mutant SNX2 markedly inhibited agonist-induced EGFR degradation, whereas internalization remained intact. In contrast, SNX1 PX domain mutants failed to effect EGFR degradation, whereas a SNX1 deletion mutant significantly inhibited receptor down-regulation. Interestingly, knockdown of SNX1 and SNX2 expression by RNA interference failed to alter agonist-induced EGFR down-regulation. Together, these findings suggest that both SNX1 and SNX2 are involved in regulating lysosomal sorting of internalized EGFR, but neither protein is essential for this process. These studies are the first to demonstrate a function for SNX2 in protein trafficking.

scholarly journals Regulation of Epidermal Growth Factor Receptor Signaling by Endocytosis and Intracellular Trafficking

2001 ◽  
Vol 12 (6) ◽  
pp. 1897-1910 ◽  
Author(s):  
Patrick Burke ◽  
Kevin Schooler ◽  
H. Steven Wiley
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Growth Factor Receptor ◽  
Egfr Signaling ◽  
Egfr Activation ◽  
Early Endosomes ◽  
Epidermal Growth ◽  
Growth Factor Receptor Signaling ◽  
Regulated Trafficking

Ligand activation of the epidermal growth factor receptor (EGFR) leads to its rapid internalization and eventual delivery to lysosomes. This process is thought to be a mechanism to attenuate signaling, but signals could potentially be generated after endocytosis. To directly evaluate EGFR signaling during receptor trafficking, we developed a technique to rapidly and selectively isolate internalized EGFR and associated molecules with the use of reversibly biotinylated anti-EGFR antibodies. In addition, we developed antibodies specific to tyrosine-phosphorylated EGFR. With the use of a combination of fluorescence imaging and affinity precipitation approaches, we evaluated the state of EGFR activation and substrate association during trafficking in epithelial cells. We found that after internalization, EGFR remained active in the early endosomes. However, receptors were inactivated before degradation, apparently due to ligand removal from endosomes. Adapter molecules, such as Shc, were associated with EGFR both at the cell surface and within endosomes. Some molecules, such as Grb2, were primarily found associated with surface EGFR, whereas others, such as Eps8, were found only with intracellular receptors. During the inactivation phase, c-Cbl became EGFR associated, consistent with its postulated role in receptor attenuation. We conclude that the association of the EGFR with different proteins is compartment specific. In addition, ligand loss is the proximal cause of EGFR inactivation. Thus, regulated trafficking could potentially influence the pattern as well as the duration of signal transduction.

scholarly journals Involvement of calpain-7 in epidermal growth factor receptor degradation via the endosomal sorting pathway

FEBS Journal ◽  
2014 ◽  
Vol 281 (16) ◽  
pp. 3642-3655 ◽  
Author(s):  
Yuki Maemoto ◽  
Yasuko Ono ◽  
Satomi Kiso ◽  
Hideki Shibata ◽  
Terunao Takahara ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Growth Factor Receptor ◽  
Endosomal Sorting ◽  
Epidermal Growth

scholarly journals c-Cbl-mediated Ubiquitinylation Is Required for Epidermal Growth Factor Receptor Exit from the Early Endosomes

2004 ◽  
Vol 279 (35) ◽  
pp. 37153-37162 ◽  
Author(s):  
Tommer Ravid ◽  
Jill M. Heidinger ◽  
Peter Gee ◽  
Elaine M. Khan ◽  
Tzipora Goldkorn
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Growth Factor Receptor ◽  
Early Endosomes ◽  
Epidermal Growth

scholarly journals Protein Kinase Cδ and Calmodulin Regulate Epidermal Growth Factor Receptor Recycling from Early Endosomes through Arp2/3 Complex and Cortactin

2008 ◽  
Vol 19 (1) ◽  
pp. 17-29 ◽  
Author(s):  
Anna Lladó ◽  
Paul Timpson ◽  
Sandra Vilà de Muga ◽  
Jemina Moretó ◽  
Albert Pol ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Protein Kinase ◽  
Epidermal Growth Factor ◽  
Intracellular Trafficking ◽  
Actin Polymerization ◽  
Growth Factor Receptor ◽  
Early Endosomes ◽  
Epidermal Growth ◽  
Protein Kinase Cδ

The intracellular trafficking of the epidermal growth factor receptor (EGFR) is regulated by a cross-talk between calmodulin (CaM) and protein kinase Cδ (PKCδ). On inhibition of CaM, PKCδ promotes the formation of enlarged early endosomes and blocks EGFR recycling and degradation. Here, we show that PKCδ impairs EGFR trafficking due to the formation of an F-actin coat surrounding early endosomes. The PKCδ-induced polymerization of actin is orchestrated by the Arp2/3 complex and requires the interaction of cortactin with PKCδ. Accordingly, inhibition of actin polymerization by using cytochalasin D or by overexpression of active cofilin, restored the normal morphology of the organelle and the recycling of EGFR. Similar results were obtained after down-regulation of cortactin and the sequestration of the Arp2/3 complex. Furthermore we demonstrate an interaction of cortactin with CaM and PKCδ, the latter being dependent on CaM inhibition. In summary, this study provides the first evidence that CaM and PKCδ organize actin dynamics in the early endosomal compartment, thereby regulating the intracellular trafficking of EGFR.

scholarly journals Protein KinaseCδ-Calmodulin Crosstalk Regulates Epidermal Growth Factor Receptor Exit from Early Endosomes

2004 ◽  
Vol 15 (11) ◽  
pp. 4877-4891 ◽  
Author(s):  
Anna Lladó ◽  
Francesc Tebar ◽  
Maria Calvo ◽  
Jemina Moretó ◽  
Alexander Sorkin ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Protein Kinase ◽  
Epidermal Growth Factor ◽  
Growth Factor Receptor ◽  
Calmodulin Antagonist ◽  
Endocytic Compartment ◽  
Early Endosomes ◽  
Epidermal Growth ◽  
In Cells

We have recently shown that calmodulin antagonist W13 interferes with the trafficking of the epidermal growth factor receptor (EGFR) and regulates the mitogen-activated protein kinase (MAPK) signaling pathway. In the present study, we demonstrate that in cells in which calmodulin is inhibited, protein kinase C (PKC) inhibitors rapidly restore EGFR and transferrin trafficking through the recycling compartment, although onward transport to the degradative pathway remains arrested. Analysis of PKC isoforms reveals that inhibition of PKCδ with rottlerin or its down-modulation by using small interfering RNA is specifically responsible for the release of the W13 blockage of EGFR trafficking from early endosomes. The use of the inhibitor Gö 6976, specific for conventional PKCs (α, β, and γ), or expression of dominant-negative forms of PKCλ, ζ, or ϵ did not restore the effects of W13. Furthermore, in cells treated with W13 and rottlerin, we observed a recovery of brefeldin A tubulation, as well as transport of dextran-fluorescein isothiocyanate toward the late endocytic compartment. These results demonstrate a specific interplay between calmodulin and PKCδ in the regulation of the morphology of and trafficking from the early endocytic compartment.

scholarly journals Gαs protein binds ubiquitin to regulate epidermal growth factor receptor endosomal sorting

2017 ◽  
Vol 114 (51) ◽  
pp. 13477-13482 ◽  
Author(s):  
Xuezhi Li ◽  
Danny Létourneau ◽  
Brian Holleran ◽  
Richard Leduc ◽  
Pierre Lavigne ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cellular Response ◽  
Growth Factor Receptor ◽  
Fine Tuning ◽  
Lysosomal Degradation ◽  
Endosomal Sorting ◽  
Epidermal Growth ◽  
G Protein Coupled

The Gαs subunit is classically involved in the signal transduction of G protein-coupled receptors (GPCRs) at the plasma membrane. Recent evidence has revealed noncanonical roles for Gαs in endosomal sorting of receptors to lysosomes. However, the mechanism of action of Gαs in this sorting step is still poorly characterized. Here, we report that Gαs interacts with ubiquitin to regulate the endosomal sorting of receptors for lysosomal degradation. We reveal that the N-terminal extremity of Gαs contains a ubiquitin-interacting motif (UIM), a sorting element usually found in the endosomal sorting complex required for transport (ESCRT) machinery responsible for sorting ubiquitinated receptors into intraluminal vesicles (ILVs) of multivesicular bodies (MVBs). Mutation of the UIM in Gαs confirmed the importance of ubiquitin interaction for the sorting of epidermal growth factor receptor (EGFR) into ILVs for lysosomal degradation. These findings demonstrate a role for Gαs as an integral component of the ubiquitin-dependent endosomal sorting machinery and highlight the dual role of Gαs in receptor trafficking and signaling for the fine-tuning of the cellular response.

A role for Hrs in endosomal sorting of ligand-stimulated and unstimulated epidermal growth factor receptor

2004 ◽  
Vol 297 (2) ◽  
pp. 380-391 ◽  
Author(s):  
Chitose Morino ◽  
Masaki Kato ◽  
Akitsugu Yamamoto ◽  
Emi Mizuno ◽  
Akira Hayakawa ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Growth Factor Receptor ◽  
Endosomal Sorting ◽  
Epidermal Growth
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