Cargando…
Synthetic control systems for high performance gene expression in mammalian cells
Tunable induction of gene expression is an essential tool in biology and biotechnology. In spite of that, current induction systems often exhibit unpredictable behavior and performance shortcomings, including high sensitivity to transactivator dosage and plasmid take-up variation, and excessive cons...
Autores principales: | , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2018
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6182142/ https://www.ncbi.nlm.nih.gov/pubmed/30203050 http://dx.doi.org/10.1093/nar/gky795 |
_version_ | 1783362498555019264 |
---|---|
author | Lillacci, Gabriele Benenson, Yaakov Khammash, Mustafa |
author_facet | Lillacci, Gabriele Benenson, Yaakov Khammash, Mustafa |
author_sort | Lillacci, Gabriele |
collection | PubMed |
description | Tunable induction of gene expression is an essential tool in biology and biotechnology. In spite of that, current induction systems often exhibit unpredictable behavior and performance shortcomings, including high sensitivity to transactivator dosage and plasmid take-up variation, and excessive consumption of cellular resources. To mitigate these limitations, we introduce here a novel family of gene expression control systems of varying complexity with significantly enhanced performance. These include: (i) an incoherent feedforward circuit that exhibits output tunability and robustness to plasmid take-up variation; (ii) a negative feedback circuit that reduces burden and provides robustness to transactivator dosage variability; and (iii) a new hybrid circuit integrating negative feedback and incoherent feedforward that combines the benefits of both. As with endogenous circuits, the complexity of our genetic controllers is not gratuitous, but is the necessary outcome of more stringent performance requirements. We demonstrate the benefits of these controllers in two applications. In a culture of CHO cells for protein manufacturing, the circuits result in up to a 2.6-fold yield improvement over a standard system. In human-induced pluripotent stem cells they enable precisely regulated expression of an otherwise poorly tolerated gene of interest, resulting in a significant increase in the viability of the transfected cells. |
format | Online Article Text |
id | pubmed-6182142 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-61821422018-10-18 Synthetic control systems for high performance gene expression in mammalian cells Lillacci, Gabriele Benenson, Yaakov Khammash, Mustafa Nucleic Acids Res Synthetic Biology and Bioengineering Tunable induction of gene expression is an essential tool in biology and biotechnology. In spite of that, current induction systems often exhibit unpredictable behavior and performance shortcomings, including high sensitivity to transactivator dosage and plasmid take-up variation, and excessive consumption of cellular resources. To mitigate these limitations, we introduce here a novel family of gene expression control systems of varying complexity with significantly enhanced performance. These include: (i) an incoherent feedforward circuit that exhibits output tunability and robustness to plasmid take-up variation; (ii) a negative feedback circuit that reduces burden and provides robustness to transactivator dosage variability; and (iii) a new hybrid circuit integrating negative feedback and incoherent feedforward that combines the benefits of both. As with endogenous circuits, the complexity of our genetic controllers is not gratuitous, but is the necessary outcome of more stringent performance requirements. We demonstrate the benefits of these controllers in two applications. In a culture of CHO cells for protein manufacturing, the circuits result in up to a 2.6-fold yield improvement over a standard system. In human-induced pluripotent stem cells they enable precisely regulated expression of an otherwise poorly tolerated gene of interest, resulting in a significant increase in the viability of the transfected cells. Oxford University Press 2018-10-12 2018-09-07 /pmc/articles/PMC6182142/ /pubmed/30203050 http://dx.doi.org/10.1093/nar/gky795 Text en © The Author(s) 2018. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com |
spellingShingle | Synthetic Biology and Bioengineering Lillacci, Gabriele Benenson, Yaakov Khammash, Mustafa Synthetic control systems for high performance gene expression in mammalian cells |
title | Synthetic control systems for high performance gene expression in mammalian cells |
title_full | Synthetic control systems for high performance gene expression in mammalian cells |
title_fullStr | Synthetic control systems for high performance gene expression in mammalian cells |
title_full_unstemmed | Synthetic control systems for high performance gene expression in mammalian cells |
title_short | Synthetic control systems for high performance gene expression in mammalian cells |
title_sort | synthetic control systems for high performance gene expression in mammalian cells |
topic | Synthetic Biology and Bioengineering |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6182142/ https://www.ncbi.nlm.nih.gov/pubmed/30203050 http://dx.doi.org/10.1093/nar/gky795 |
work_keys_str_mv | AT lillaccigabriele syntheticcontrolsystemsforhighperformancegeneexpressioninmammaliancells AT benensonyaakov syntheticcontrolsystemsforhighperformancegeneexpressioninmammaliancells AT khammashmustafa syntheticcontrolsystemsforhighperformancegeneexpressioninmammaliancells |