Endocytosis of functional epidermal growth factor receptor-green fluorescent protein chimera - PubMed (original) (raw)
. 1998 Dec 25;273(52):35000-7.
doi: 10.1074/jbc.273.52.35000.
Affiliations
- PMID: 9857032
- DOI: 10.1074/jbc.273.52.35000
Free article
Endocytosis of functional epidermal growth factor receptor-green fluorescent protein chimera
R E Carter et al. J Biol Chem. 1998.
Free article
Abstract
A chimera of the epidermal growth factor receptor (EGFR) and green fluorescent protein (GFP) has been engineered by fusing GFP to the carboxyl terminus of EGFR. Data are provided to demonstrate that the GFP moiety does not affect the expected functioning of EGFR. EGFR-GFP becomes phosphorylated at tyrosine residues in response to EGF and is capable of phosphorylating endogenous substrates and initiating signaling cascades. EGF-dependent association of the chimeric receptor with the clathrin adaptor protein AP-2, involved in endocytosis, and with Shc adaptor protein, which binds in close proximity to the fusion point, is not affected by the GFP moiety. Receptor down-regulation and internalization occur at rates similar to those in cells expressing wild-type EGFR. Western blot analysis reveals that lysosomal degradation of EGFR-GFP proceeds from the extracellular domain and that GFP is not preferentially cleaved. Time-dependent co-localization of EGFR-GFP and Texas Red-conjugated EGF in living cells using digital deconvolution microscopy demonstrates the trafficking of ligand-receptor complexes through the early and multivesicular endosomes followed by segregation of the ligand and receptor at the late stages of endocytosis. Time-lapse optical analysis of the early stages of endocytosis reveals localization of EGFR-GFP in the tubular-vesicular endosomal compartments. Rapid dynamics of membrane movement and fusion within these compartments were observed. This approach and the fidelity of the biochemical properties of the EGFR-GFP demonstrate that real-time visualization of trafficking and protein interactions of tyrosine kinase receptors in the presence or absence of the ligand are feasible.
Similar articles
- Trafficking of TrkA-green fluorescent protein chimerae during nerve growth factor-induced differentiation.
Jullien J, Guili V, Derrington EA, Darlix JL, Reichardt LF, Rudkin BB. Jullien J, et al. J Biol Chem. 2003 Mar 7;278(10):8706-16. doi: 10.1074/jbc.M202401200. Epub 2002 Nov 15. J Biol Chem. 2003. PMID: 12438306 Free PMC article. - Live-cell fluorescence imaging reveals high stoichiometry of Grb2 binding to the EGF receptor sustained during endocytosis.
Fortian A, Sorkin A. Fortian A, et al. J Cell Sci. 2014 Jan 15;127(Pt 2):432-44. doi: 10.1242/jcs.137786. Epub 2013 Nov 20. J Cell Sci. 2014. PMID: 24259669 Free PMC article. - Adaptor protein Shc undergoes translocation and mediates up-regulation of the tyrosine kinase c-Src in EGF-stimulated A431 cells.
Sato K, Kimoto M, Kakumoto M, Horiuchi D, Iwasaki T, Tokmakov AA, Fukami Y. Sato K, et al. Genes Cells. 2000 Sep;5(9):749-64. doi: 10.1046/j.1365-2443.2000.00358.x. Genes Cells. 2000. PMID: 10971656 - Endocytosis and intracellular trafficking of ErbBs.
Sorkin A, Goh LK. Sorkin A, et al. Exp Cell Res. 2008 Oct 15;314(17):3093-106. doi: 10.1016/j.yexcr.2008.08.013. Epub 2008 Aug 28. Exp Cell Res. 2008. PMID: 18793634 Free PMC article. Review. - Internalization of the epidermal growth factor receptor: role in signalling.
Sorkin A. Sorkin A. Biochem Soc Trans. 2001 Aug;29(Pt 4):480-4. doi: 10.1042/bst0290480. Biochem Soc Trans. 2001. PMID: 11498013 Review.
Cited by
- Sushi domain-containing protein 4 binds to epithelial growth factor receptor and initiates autophagy in an EGFR phosphorylation independent manner.
Papadakos KS, Ekström A, Slipek P, Skourti E, Reid S, Pietras K, Blom AM. Papadakos KS, et al. J Exp Clin Cancer Res. 2022 Dec 29;41(1):363. doi: 10.1186/s13046-022-02565-1. J Exp Clin Cancer Res. 2022. PMID: 36578014 Free PMC article. - N6-Isopentenyladenosine Impairs Mitochondrial Metabolism through Inhibition of EGFR Translocation on Mitochondria in Glioblastoma Cells.
Pagano C, Coppola L, Navarra G, Avilia G, Bruzzaniti S, Piemonte E, Galgani M, Della Monica R, Chiariotti L, Cuomo M, Buonaiuto M, Torelli G, Caiazzo P, Laezza C, Bifulco M. Pagano C, et al. Cancers (Basel). 2022 Dec 8;14(24):6044. doi: 10.3390/cancers14246044. Cancers (Basel). 2022. PMID: 36551529 Free PMC article. - Benzimidazole-linked pyrazolo[1,5-a]pyrimidine conjugates: synthesis and detail evaluation as potential anticancer agents.
Bagul C, Rao GK, Veena I, Kulkarni R, Tamboli JR, Akunuri R, Shaik SP, Pal-Bhadra M, Kamal A. Bagul C, et al. Mol Divers. 2023 Jun;27(3):1185-1202. doi: 10.1007/s11030-022-10481-x. Epub 2022 Sep 17. Mol Divers. 2023. PMID: 36114912 - A Multiscale Molecular Dynamic Analysis Reveals the Effect of Sialylation on EGFR Clustering in a CRISPR/Cas9-Derived Model.
Leong SK, Hsiao JC, Shie JJ. Leong SK, et al. Int J Mol Sci. 2022 Aug 6;23(15):8754. doi: 10.3390/ijms23158754. Int J Mol Sci. 2022. PMID: 35955894 Free PMC article.
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources
Research Materials
Miscellaneous