Engineering of non-model eukaryotes for bioenergy and biochemical production

Ploessl, Deon; Zhao, Yuxin; Shao, Zengyi

doi:10.1016/j.copbio.2022.102869

Cited by 10 publications

(4 citation statements)

References 78 publications

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“…This represents a genomeinformed advancement within the field of broad-host-range biodesign which aims to lessen our reliance on model organisms so that we can better understand diverse microbial behaviors and use them in the blueprints of biodesign. The number of assimilated microbes available for use as industrial biotechnology platforms and biodesign engineering is increasing steadily [58][59][60] , heralding the advancement towards broad-…”

Section: Discussionmentioning

confidence: 99%

“…This represents a genome-informed advancement within the field of broad-host-range biodesign which aims to lessen our reliance on model organisms so that we can better understand diverse microbial behaviors and use them in the blueprints of biodesign. The number of assimilated microbes available for use as 30 industrial biotechnology platforms and biodesign engineering is increasing steadily [58][59][60] , heralding the advancement towards broad-host-range synthetic biology where synthetic biologists must explore not only the design space of genetic parts, but also the design space of host-chassis in order to optimize their engineered systems. As the field of biodesign progresses towards this new era, the constraints imposed by the chassis-effect will only become more relevant.…”

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confidence: 99%

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Pangenomic landscapes shape the genetic circuit performance in aStutzerimonasbiodesign toolkit

Chan,

Bernstein

2024

Preprint

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Implementation of identical biodesign strategies into different species often results in different performance, a process called thechassis effect. Our current understanding of how cellular host context underpins its ability to be engineered is lacking and closing this knowledge gap will greatly improve the rational design of microorganisms. Here, we combined global differential gene expression analysis and pangenomics to uncover how genome structure and function relates to the observed chassis effect of an engineered genetic inverter device operating in six closely relatedStutzerimonashosts. The differential expression of the core genome, gene clusters shared between all hosts, was found to be the main source of significant concordance to the observed device performance, whereas specialty genes from respective accessory genomes were not significant. A data-driven investigation revealed that genes related to denitrification and efflux pumps were among the most differentially expressed gene clusters in response to the engineered device. This study establishes that the effectiveness of synthetic gene circuits can be traced along differences in closely related microbial hosts that each represent unique hardware options for biodesign.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Pangenomic landscapes shape the genetic circuit performance in aStutzerimonasbiodesign toolkit

Chan,

Bernstein

2024

Preprint

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“…The field of biology has been revolutionized by the discovery of adaptive immune systems encoded by clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPRassociated (Cas) genes, and the subsequent harnessing of CRISPR-Cas systems for genome engineering. In particular, CRISPR-Cas based genetic manipulations have been applied to several model (i.e., E. coli) and non-model organisms across all domains of life with high efficiency [1][2][3][4] which has greatly aided our understanding of the biology underlying these organisms. CRISPR-Cas systems encode a diverse repitoire of RNA-guided CRISPR-associated (Cas) effector nucleases that perform interference on invading mobile genetic elements.…”

Section: Introductionmentioning

confidence: 99%

Bacterial genome engineering using CRISPR RNA-guided transposases

Gelsinger

Klompe

et al. 2023

Preprint

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CRISPR-associated transposons (CASTs) have the potential to transform the technology landscape for kilobase-scale genome engineering, by virtue of their ability to integrate large genetic payloads with high accuracy, easy programmability, and no requirement for homologous recombination machinery. These transposons encode efficient, CRISPR RNA-guided transposases that execute genomic insertions in E. coli at efficiencies approaching ~100%, generate multiplexed edits when programmed with multiple guides, and function robustly in diverse Gram-negative bacterial species. Here we present a detailed protocol for engineering bacterial genomes using CAST systems, including guidelines on the available homologs and vectors, customization of guide RNAs and DNA payloads, selection of common delivery methods, and genotypic analysis of integration events. We further describe a computational crRNA design algorithm to avoid potential off-targets and CRISPR array cloning pipeline for DNA insertion multiplexing. Starting from available plasmid constructs, the isolation of clonal strains containing a novel genomic integration event-of-interest can be achieved in 1 week using standard molecular biology techniques.

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“…In the post-CRISPR era, rapid advancement in the exploration of unique microbial species challenges the status of E. coli and S. cerevisiae as classical microbial cell factories 20 . Many nonconventional microorganisms are of great interest in industrial applications (e.g., high temperature and low pH) due to their innate traits with respect to their unique desirable physiology, metabolism, biosynthesis, and fermentation capacity, which are acquired through a long-term natural evolution in specific environments 21 , 22 .…”

Section: Introductionmentioning

confidence: 99%

Xylose and shikimate transporters facilitates microbial consortium as a chassis for benzylisoquinoline alkaloid production

Gao,

Zhao,

Yao

et al. 2023

Nat Commun

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Plant-sourced aromatic amino acid (AAA) derivatives are a vast group of compounds with broad applications. Here, we present the development of a yeast consortium for efficient production of (S)-norcoclaurine, the key precursor for benzylisoquinoline alkaloid biosynthesis. A xylose transporter enables the concurrent mixed-sugar utilization in Scheffersomyces stipitis, which plays a crucial role in enhancing the flux entering the highly regulated shikimate pathway located upstream of AAA biosynthesis. Two quinate permeases isolated from Aspergillus niger facilitates shikimate translocation to the co-cultured Saccharomyces cerevisiae that converts shikimate to (S)-norcoclaurine, resulting in the maximal titer (11.5 mg/L), nearly 110-fold higher than the titer reported for an S. cerevisiae monoculture. Our findings magnify the potential of microbial consortium platforms for the economical de novo synthesis of complex compounds, where pathway modularization and compartmentalization in distinct specialty strains enable effective fine-tuning of long biosynthetic pathways and diminish intermediate buildup, thereby leading to increases in production.

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Engineering of non-model eukaryotes for bioenergy and biochemical production

Cited by 10 publications

References 78 publications

Pangenomic landscapes shape the genetic circuit performance in aStutzerimonasbiodesign toolkit

Pangenomic landscapes shape the genetic circuit performance in aStutzerimonasbiodesign toolkit

Bacterial genome engineering using CRISPR RNA-guided transposases

Xylose and shikimate transporters facilitates microbial consortium as a chassis for benzylisoquinoline alkaloid production

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