Activation of the EPO-R [EPO (erythropoietin) receptor] by its ligand EPO promotes erythropoiesis. Low cell surface EPO-R levels are traditionally attributed to inefficient folding mediated by the receptor extracellular domain. In the present study, we addressed the role of the EPO-R intracellular domain in exit from the ER (endoplasmic reticulum) and surface expression. A fusion protein between the thermo-reversible folding mutant of VSVG (vesicular-stomatitis-virus glycoprotein) (VSVGtsO45) and the EPO-R cytosolic domain [VSVG-WT (wild-type)] displayed delayed intracellular trafficking as compared with the parental VSVGtsO45, suggesting that the EPO-R cytosolic domain can hamper ER exit. Although NPXY-based motifs were originally associated with clathrin binding and endocytosis, they may also function in other contexts of the secretory pathway. A fusion protein between VSVGtsO45 and the cytosolic portion of EPO-R containing an NPVY insert (VSVG-NPVY) displayed enhanced glycan maturation and surface expression as compared with VSVG-WT. Notably, the NPVY insert also conferred improved maturation and augmented cell surface EPO-R. Our findings highlight three major concepts: (i) the EPO-R cytosolic domain is involved in ER exit of the receptor. (ii) Sequence motifs that participate in endocytosis can also modulate transport along the secretory pathway. (iii) VSVG-fusion proteins may be employed to screen for intracellular sequences that regulate transport.
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Research Article|
February 12 2008
A transplanted NPVY sequence in the cytosolic domain of the erythropoietin receptor enhances maturation
Tal Nahari;
Tal Nahari
*Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel-Aviv University, Ramat-Aviv 69978, Israel
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Eran Barzilay;
Eran Barzilay
*Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel-Aviv University, Ramat-Aviv 69978, Israel
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Koret Hirschberg;
Koret Hirschberg
†Department of Pathology, Sackler Faculty of Medicine, Tel-Aviv University, Ramat-Aviv 69978, Israel
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Drorit Neumann
Drorit Neumann
1
*Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel-Aviv University, Ramat-Aviv 69978, Israel
1To whom correspondence should be addressed (email Histo6@post.tau.ac.il).
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Publisher: Portland Press Ltd
Received:
September 20 2007
Revision Received:
November 05 2007
Accepted:
November 12 2007
Accepted Manuscript online:
November 12 2007
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem J (2008) 410 (2): 409–416.
Article history
Received:
September 20 2007
Revision Received:
November 05 2007
Accepted:
November 12 2007
Accepted Manuscript online:
November 12 2007
Citation
Tal Nahari, Eran Barzilay, Koret Hirschberg, Drorit Neumann; A transplanted NPVY sequence in the cytosolic domain of the erythropoietin receptor enhances maturation. Biochem J 1 March 2008; 410 (2): 409–416. doi: https://doi.org/10.1042/BJ20071297
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