Application of combinatorial optimization strategies in synthetic biology

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Abstract

In the first wave of synthetic biology, genetic elements, combined into simple circuits, are used to control individual cellular functions. In the second wave of synthetic biology, the simple circuits, combined into complex circuits, form systems-level functions. However, efforts to construct complex circuits are often impeded by our limited knowledge of the optimal combination of individual circuits. For example, a fundamental question in most metabolic engineering projects is the optimal level of enzymes for maximizing the output. To address this point, combinatorial optimization approaches have been established, allowing automatic optimization without prior knowledge of the best combination of expression levels of individual genes. This review focuses on current combinatorial optimization methods and emerging technologies facilitating their applications.

Abstract

Our efforts to build complex synthetic biology circuits are impeded by limited knowledge of optimal combinations. In this review, the authors consider current combinatorial methods and look to emerging technologies.

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Programming cells by multiplex genome engineering and accelerated evolution.

Harris Wang, Farren J. Isaacs, Peter A Carr … (2009)

The breadth of genomic diversity found among organisms in nature allows populations to adapt to diverse environments. However, genomic diversity is difficult to generate in the laboratory and new phenotypes do not easily arise on practical timescales. Although in vitro and directed evolution methods have created genetic variants with usefully altered phenotypes, these methods are limited to laborious and serial manipulation of single genes and are not used for parallel and continuous directed evolution of gene networks or genomes. Here, we describe multiplex automated genome engineering (MAGE) for large-scale programming and evolution of cells. MAGE simultaneously targets many locations on the chromosome for modification in a single cell or across a population of cells, thus producing combinatorial genomic diversity. Because the process is cyclical and scalable, we constructed prototype devices that automate the MAGE technology to facilitate rapid and continuous generation of a diverse set of genetic changes (mismatches, insertions, deletions). We applied MAGE to optimize the 1-deoxy-D-xylulose-5-phosphate (DXP) biosynthesis pathway in Escherichia coli to overproduce the industrially important isoprenoid lycopene. Twenty-four genetic components in the DXP pathway were modified simultaneously using a complex pool of synthetic DNA, creating over 4.3 billion combinatorial genomic variants per day. We isolated variants with more than fivefold increase in lycopene production within 3 days, a significant improvement over existing metabolic engineering techniques. Our multiplex approach embraces engineering in the context of evolution by expediting the design and evolution of organisms with new and improved properties.

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Optical Control of Mammalian Endogenous Transcription and Epigenetic States

Silvana Konermann, Mark Brigham, Alexandro E Trevino … (2013)

The dynamic nature of gene expression enables cellular programming, homeostasis, and environmental adaptation in living systems. Dissection of causal gene functions in cellular and organismal processes therefore necessitates approaches that enable spatially and temporally precise modulation of gene expression. Recently, a variety of microbial and plant-derived light-sensitive proteins have been engineered as optogenetic actuators, enabling high precision spatiotemporal control of many cellular functions 1-11 . However, versatile and robust technologies that enable optical modulation of transcription in the mammalian endogenous genome remain elusive. Here, we describe the development of Light-Inducible Transcriptional Effectors (LITEs), an optogenetic two-hybrid system integrating the customizable TALE DNA-binding domain 12-14 with the light-sensitive cryptochrome 2 protein and its interacting partner CIB1 from Arabidopsis thaliana. LITEs do not require additional exogenous chemical co-factors, are easily customized to target many endogenous genomic loci, and can be activated within minutes with reversibility 3,4,6,7,15 . LITEs can be packaged into viral vectors and genetically targeted to probe specific cell populations. We have applied this system in primary mouse neurons, as well as in the brain of awake mice in vivo to mediate reversible modulation of mammalian endogenous gene expression as well as targeted epigenetic chromatin modifications. The LITE system establishes a novel mode of optogenetic control of endogenous cellular processes and enables direct testing of the causal roles of genetic and epigenetic regulation in normal biological processes and disease states.

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De novo production of the plant-derived alkaloid strictosidine in yeast.

Stephanie Brown, Marc Clastre, Vincent Courdavault … (2015)

The monoterpene indole alkaloids are a large group of plant-derived specialized metabolites, many of which have valuable pharmaceutical or biological activity. There are ∼3,000 monoterpene indole alkaloids produced by thousands of plant species in numerous families. The diverse chemical structures found in this metabolite class originate from strictosidine, which is the last common biosynthetic intermediate for all monoterpene indole alkaloid enzymatic pathways. Reconstitution of biosynthetic pathways in a heterologous host is a promising strategy for rapid and inexpensive production of complex molecules that are found in plants. Here, we demonstrate how strictosidine can be produced de novo in a Saccharomyces cerevisiae host from 14 known monoterpene indole alkaloid pathway genes, along with an additional seven genes and three gene deletions that enhance secondary metabolism. This system provides an important resource for developing the production of more complex plant-derived alkaloids, engineering of nonnatural derivatives, identification of bottlenecks in monoterpene indole alkaloid biosynthesis, and discovery of new pathway genes in a convenient yeast host.

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Author and article information

Contributors

Gita Naseri:

ORCID: http://orcid.org/0000-0002-4630-3141

gita.naseri@hu-berlin.de

Mattheos A. G. Koffas:

ORCID: http://orcid.org/0000-0002-1405-0565

koffam@rpi.edu

Journal

Journal ID (nlm-ta): Nat Commun

Journal ID (iso-abbrev): Nat Commun

Title: Nature Communications

Publisher: Nature Publishing Group UK (London )

ISSN (Electronic): 2041-1723

Publication date (Electronic): 15 May 2020

Publication date PMC-release: 15 May 2020

Publication date Collection: 2020

Volume: 11

Electronic Location Identifier: 2446

Affiliations

[1 ]ISNI 0000 0001 2248 7639, GRID grid.7468.d, Institut für Chemie, Humboldt Universität zu Berlin, ; 12489 Berlin, Germany

[2 ]ISNI 0000 0001 2160 9198, GRID grid.33647.35, Center for Biotechnology, , Rensselaer Polytechnic Institute, ; Troy, NY USA

[3 ]ISNI 0000 0001 2160 9198, GRID grid.33647.35, Department of Chemical and Biological Engineering, , Rensselaer Polytechnic Institute, ; Troy, NY USA

[4 ]ISNI 0000 0001 2160 9198, GRID grid.33647.35, Department of Biological Sciences, , Rensselaer Polytechnic Institute, ; Troy, NY USA

Author information

Gita Naseri http://orcid.org/0000-0002-4630-3141

Mattheos A. G. Koffas http://orcid.org/0000-0002-1405-0565

Article

Publisher ID: 16175

DOI: 10.1038/s41467-020-16175-y

PMC ID: 7229011

PubMed ID: 32415065

SO-VID: 8dad744a-1d09-43eb-b85e-699e935edde3

License:

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

History

Date received : 26 January 2019

Date accepted : 15 April 2020

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ScienceOpen disciplines: Uncategorized

Keywords: metabolic engineering,synthetic biology,machine learning,crispr-cas9 genome editing

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ScienceOpen disciplines: Uncategorized

Keywords: metabolic engineering, synthetic biology, machine learning, crispr-cas9 genome editing

Application of combinatorial optimization strategies in synthetic biology

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CRISPR/Cas9 editing in human blood

Most cited references 100

Programming cells by multiplex genome engineering and accelerated evolution.

Optical Control of Mammalian Endogenous Transcription and Epigenetic States

De novo production of the plant-derived alkaloid strictosidine in yeast.

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