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Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein

Lentiviral vectors (LVs) are a highly valuable tool for gene transfer currently exploited in basic, applied, and clinical studies. Their optimization is therefore very important for the field of vectorology and gene therapy. A key molecule for LV function is the envelope because it guides cell entry...

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Autores principales: Zucchelli, Eleonora, Pema, Monika, Stornaiuolo, Anna, Piovan, Claudia, Scavullo, Cinzia, Giuliani, Erica, Bossi, Sergio, Corna, Stefano, Asperti, Claudia, Bordignon, Claudio, Rizzardi, Gian-Paolo, Bovolenta, Chiara
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society of Gene & Cell Therapy 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5363313/
https://www.ncbi.nlm.nih.gov/pubmed/28344996
http://dx.doi.org/10.1016/j.omtm.2017.01.002
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author Zucchelli, Eleonora
Pema, Monika
Stornaiuolo, Anna
Piovan, Claudia
Scavullo, Cinzia
Giuliani, Erica
Bossi, Sergio
Corna, Stefano
Asperti, Claudia
Bordignon, Claudio
Rizzardi, Gian-Paolo
Bovolenta, Chiara
author_facet Zucchelli, Eleonora
Pema, Monika
Stornaiuolo, Anna
Piovan, Claudia
Scavullo, Cinzia
Giuliani, Erica
Bossi, Sergio
Corna, Stefano
Asperti, Claudia
Bordignon, Claudio
Rizzardi, Gian-Paolo
Bovolenta, Chiara
author_sort Zucchelli, Eleonora
collection PubMed
description Lentiviral vectors (LVs) are a highly valuable tool for gene transfer currently exploited in basic, applied, and clinical studies. Their optimization is therefore very important for the field of vectorology and gene therapy. A key molecule for LV function is the envelope because it guides cell entry. The most commonly used in transiently produced LVs is the vesicular stomatitis virus glycoprotein (VSV-G) envelope, whose continuous expression is, however, toxic for stable LV producer cells. In contrast, the feline endogenous retroviral RD114-TR envelope is suitable for stable LV manufacturing, being well tolerated by producer cells under constitutive expression. We have previously reported successful, transient and stable production of LVs pseudotyped with RD114-TR for good transduction of T lymphocytes and CD34(+) cells. To further improve RD114-TR-pseudotyped LV cell entry by increasing envelope expression, we codon-optimized the RD114-TR open reading frame (ORF). Here we show that, despite the RD114-TRco precursor being produced at a higher level than the wild-type counterpart, it is unexpectedly not duly glycosylated, exported to the cytosol, and processed. Correct cleavage of the precursor in the functional surface and transmembrane subunits is prevented in vivo, and, consequently, the unprocessed precursor is incorporated into LVs, making them inactive.
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spelling pubmed-53633132017-03-24 Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein Zucchelli, Eleonora Pema, Monika Stornaiuolo, Anna Piovan, Claudia Scavullo, Cinzia Giuliani, Erica Bossi, Sergio Corna, Stefano Asperti, Claudia Bordignon, Claudio Rizzardi, Gian-Paolo Bovolenta, Chiara Mol Ther Methods Clin Dev Original Article Lentiviral vectors (LVs) are a highly valuable tool for gene transfer currently exploited in basic, applied, and clinical studies. Their optimization is therefore very important for the field of vectorology and gene therapy. A key molecule for LV function is the envelope because it guides cell entry. The most commonly used in transiently produced LVs is the vesicular stomatitis virus glycoprotein (VSV-G) envelope, whose continuous expression is, however, toxic for stable LV producer cells. In contrast, the feline endogenous retroviral RD114-TR envelope is suitable for stable LV manufacturing, being well tolerated by producer cells under constitutive expression. We have previously reported successful, transient and stable production of LVs pseudotyped with RD114-TR for good transduction of T lymphocytes and CD34(+) cells. To further improve RD114-TR-pseudotyped LV cell entry by increasing envelope expression, we codon-optimized the RD114-TR open reading frame (ORF). Here we show that, despite the RD114-TRco precursor being produced at a higher level than the wild-type counterpart, it is unexpectedly not duly glycosylated, exported to the cytosol, and processed. Correct cleavage of the precursor in the functional surface and transmembrane subunits is prevented in vivo, and, consequently, the unprocessed precursor is incorporated into LVs, making them inactive. American Society of Gene & Cell Therapy 2017-01-11 /pmc/articles/PMC5363313/ /pubmed/28344996 http://dx.doi.org/10.1016/j.omtm.2017.01.002 Text en © 2017 The Authors http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Original Article
Zucchelli, Eleonora
Pema, Monika
Stornaiuolo, Anna
Piovan, Claudia
Scavullo, Cinzia
Giuliani, Erica
Bossi, Sergio
Corna, Stefano
Asperti, Claudia
Bordignon, Claudio
Rizzardi, Gian-Paolo
Bovolenta, Chiara
Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein
title Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein
title_full Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein
title_fullStr Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein
title_full_unstemmed Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein
title_short Codon Optimization Leads to Functional Impairment of RD114-TR Envelope Glycoprotein
title_sort codon optimization leads to functional impairment of rd114-tr envelope glycoprotein
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5363313/
https://www.ncbi.nlm.nih.gov/pubmed/28344996
http://dx.doi.org/10.1016/j.omtm.2017.01.002
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