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Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells

EGF complexed to fluorescent photostable quantum dots by biotin-streptavidin system (bEGF-savQD) is attractive for both the basic research and therapeutic application such as targeted drug delivery in EGF-receptor (EGFR) expressing cancers. However, compared to native EGF, the large size of QD and i...

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Autores principales: Salova, Anna V., Belyaeva, Tatiana N., Leontieva, Ekaterina A., Zlobina, Maria V., Kharchenko, Marianna V., Kornilova, Elena S.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Impact Journals LLC 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4868738/
https://www.ncbi.nlm.nih.gov/pubmed/26716513
http://dx.doi.org/10.18632/oncotarget.6720
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author Salova, Anna V.
Belyaeva, Tatiana N.
Leontieva, Ekaterina A.
Zlobina, Maria V.
Kharchenko, Marianna V.
Kornilova, Elena S.
author_facet Salova, Anna V.
Belyaeva, Tatiana N.
Leontieva, Ekaterina A.
Zlobina, Maria V.
Kharchenko, Marianna V.
Kornilova, Elena S.
author_sort Salova, Anna V.
collection PubMed
description EGF complexed to fluorescent photostable quantum dots by biotin-streptavidin system (bEGF-savQD) is attractive for both the basic research and therapeutic application such as targeted drug delivery in EGF-receptor (EGFR) expressing cancers. However, compared to native EGF, the large size of QD and its quasi-multivalency can have unpredictable effects on EGFR endocytosis changing the internalization portal and/or endosomal processing tightly bound to EGF signaling. We have found that bEGF-savQDs enter HeLa cells via the temperature-dependent clathrin-mediated EGF-receptor-specific pathway characteristic for native EGF. We also found that EGF-to-QD concentration ratios used for the complex preparation and the level of EGF receptor expression affect the number and integral densities of the formed endosomes. So, at EGF-to-QD ratio from 4:1 to 12:1 (at nanomolar bEGF concentrations) on average 100 bright endosomes per HeLa cell were formed 15 min after the complex addition, while 1:1 ratio resulted in formation of very few dim endosomes. However, in A431 cells overexpressing EGFR 1:1 ratio was effective. Using dynamin inhibition and Na-acidic washout we showed that bEGF-savQDs bind surface receptors and enter clathrin-coated pits slower than the same ligands without QD. Yet, the bEGF-savQD demonstrated similar to native EGF and bEGF-savCy3 co-localization dynamics with tethering protein EEA1 and HRS, the key component of sorting ESCRT0 complex. In conclusion, our comparative study reveals that in respect to entrapment into coated pits, endosomal recruitment, endosome fusions, and the initial steps of endosomal maturation, bEGF-savQD behaves like native EGF and QD implementation does not affect these important events.
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spelling pubmed-48687382016-05-20 Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells Salova, Anna V. Belyaeva, Tatiana N. Leontieva, Ekaterina A. Zlobina, Maria V. Kharchenko, Marianna V. Kornilova, Elena S. Oncotarget Research Paper EGF complexed to fluorescent photostable quantum dots by biotin-streptavidin system (bEGF-savQD) is attractive for both the basic research and therapeutic application such as targeted drug delivery in EGF-receptor (EGFR) expressing cancers. However, compared to native EGF, the large size of QD and its quasi-multivalency can have unpredictable effects on EGFR endocytosis changing the internalization portal and/or endosomal processing tightly bound to EGF signaling. We have found that bEGF-savQDs enter HeLa cells via the temperature-dependent clathrin-mediated EGF-receptor-specific pathway characteristic for native EGF. We also found that EGF-to-QD concentration ratios used for the complex preparation and the level of EGF receptor expression affect the number and integral densities of the formed endosomes. So, at EGF-to-QD ratio from 4:1 to 12:1 (at nanomolar bEGF concentrations) on average 100 bright endosomes per HeLa cell were formed 15 min after the complex addition, while 1:1 ratio resulted in formation of very few dim endosomes. However, in A431 cells overexpressing EGFR 1:1 ratio was effective. Using dynamin inhibition and Na-acidic washout we showed that bEGF-savQDs bind surface receptors and enter clathrin-coated pits slower than the same ligands without QD. Yet, the bEGF-savQD demonstrated similar to native EGF and bEGF-savCy3 co-localization dynamics with tethering protein EEA1 and HRS, the key component of sorting ESCRT0 complex. In conclusion, our comparative study reveals that in respect to entrapment into coated pits, endosomal recruitment, endosome fusions, and the initial steps of endosomal maturation, bEGF-savQD behaves like native EGF and QD implementation does not affect these important events. Impact Journals LLC 2015-12-22 /pmc/articles/PMC4868738/ /pubmed/26716513 http://dx.doi.org/10.18632/oncotarget.6720 Text en Copyright: © 2016 Salova et al. http://creativecommons.org/licenses/by/2.5/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Paper
Salova, Anna V.
Belyaeva, Tatiana N.
Leontieva, Ekaterina A.
Zlobina, Maria V.
Kharchenko, Marianna V.
Kornilova, Elena S.
Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells
title Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells
title_full Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells
title_fullStr Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells
title_full_unstemmed Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells
title_short Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells
title_sort quantum dots implementation as a label for analysis of early stages of egf receptor endocytosis: a comparative study on cultured cells
topic Research Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4868738/
https://www.ncbi.nlm.nih.gov/pubmed/26716513
http://dx.doi.org/10.18632/oncotarget.6720
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