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Current advances of biocontainment strategy in synthetic biology

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Chinese Journal of Chemical Engineering 2023年第4期56卷 141-151页

作者： Xueying Zhu Zhaoyang Zhang Bin Jia yingjin yuan Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin 300072China

Synthetic biotechnology has led to the widespread application of genetically modified organisms(GMOs)in biochemistry, bioenergy, and therapy. However, the uncontrolled spread of GMOs may lead to genetic contamination by horizontal gene transfer, resulting in unpredictable biosafety risks. To deal with these challenges, many effective methods have been developed for biocontainment. In this article, we summarize and discuss recent advances in biocontainment strategies from three aspects: DNA replication, transcriptional regulation, and protein translation. We also briefly introduce the efforts in the biocontainment convention, such as the recent publication of the Tianjin Biosecurity Guidelines for the Code of Conduct for Scientists.

关键词： Genetically modified organisms(GMOs) Biocontainment Biosecurity Synthetic biology Molecular biology Biotechnology

Directed yeast genome evolution by controlled introduction of trans-chromosomic structural variations

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Science China(Life Sciences) 2022年第9期65卷 1703-1717页

作者： Bin Jia Jin Jin Mingzhe Han Bingzhi Li yingjin yuan Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin 300072China

Naturally occurring structural variations(SVs)are a considerable source of genomic variation that can reshape the 3D architecture of chromosomes.Controllable methods aimed at introducing the complex SVs and their related molecular mechanisms have remained farfetched.In this study,an SV-prone yeast strain was developed using Synthetic Chromosome Rearrangement and Modification by LoxP-mediated Evolution(SCRaMbLE)technology with two synthetic chromosomes,namely synV and synX.The biosynthesis of astaxanthin is used as a readout and a proof of concept for the application of SVs in industries.Our findings showed that complex SVs,including a pericentric inversion and a trans-chromosome translocation between synV and synX,resulted in two neo-chromosomes and a 2.7-fold yield of astaxanthin.Also,genetic targets were mapped,which resulted in a higher astaxanthin yield,thus demonstrating the SVs’ability to reorganize genetic information along the chromosomes.The rational design of trans-chromosome translocation and pericentric inversion enabled precise induction of these phenomena.Collectively,this study provides an effective tool to not only accelerate the directed genome evolution but also to reveal the mechanistic insight of complex SVs for altering phenotypes.

关键词： directed genome evolution structural variations SCRaMbLE synthetic genome synthetic biology

Design and construction of microbial cell factories based on systems biology

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Synthetic and Systems Biotechnology 2023年第1期8卷 176-185页

作者： Wenlong Yan Zhibei Cao Mingzhu Ding yingjin yuan Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin300072China Collaborative Innovation Center of Chemical Science and Engineering(Tianjin) Tianjin UniversityTianjin300072China

Environmental sustainability is an increasingly important issue in industry.As an environmentally friendly and sustainable way,constructing microbial cell factories to produce all kinds of valuable products has attracted more and more attention.In the process of constructing microbial cell factories,systems biology plays a crucial role.This review summarizes the recent applications of systems biology in the design and construction of microbial cell factories from four perspectives,including functional genes/enzymes discovery,bottleneck pathways identification,strains tolerance improvement and design and construction of synthetic microbial consortia.Systems biology tools can be employed to identify functional genes/enzymes involved in the biosynthetic pathways of products.These discovered genes are introduced into appropriate chassis strains to build engineering microorganisms capable of producing products.Subsequently,systems biology tools are used to identify bottleneck pathways,improve strains tolerance and guide design and construction of synthetic microbial consortia,resulting in increasing the yield of engineered strains and constructing microbial cell factories successfully.

关键词： Microbial cell factories Systems biology Functional genes/enzymes discovery Bottleneck pathways Strains tolerance Synthetic microbial consortia

Enhancing whole yeast genome rearrangements through multiple LoxP sym sequences

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Science China Life Sciences 2024年

作者： Bing-Zhi Li Ying-Jin yuan Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education) School of Chemical Engineering and Technology Tianjin University Frontiers Research Institute for Synthetic Biology Tianjin University

Genomic rearrangements play a significant role in disease, evolution, and tumorigenesis(Mani and Chinnaiyan,2010). While single nucleotide polymorphisms(SNPs) and small insertions/deletions contribute substantially to genetic variation, discerning the independent impact of structural variations poses a challenge. Additionally, experimental induction and control of structural variations present difficulties that hinder further investigation and exploitation.

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Construction of synthetic microbial consortia for 2-keto-L-gulonic acid biosynthesis

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Synthetic and Systems Biotechnology 2022年第1期7卷 481-489页

作者： Yan Wang Hengchang Li Yu Liu Mengyu Zhou Mingzhu Ding yingjin yuan Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin300072China Collaborative Innovation Center of Chemical Science and Engineering(Tianjin) Tianjin UniversityTianjin300072China

Currently,the establishment of synthetic microbial consortia with rational strategies has gained extensive attention,becoming one of the important frontiers of synthetic biology.Systems biology can offer insights into the design and construction of synthetic microbial consortia.Taking the high-efficiency production of 2-keto-L-gulonic acid(2-KLG)as an example,we constructed a synthetic microbial consortium“Saccharomyces cerevisiae-Ketogulonigenium vulgare”based on systems biology analysis.In the consortium,K.vulgare was the 2-KLG pro-ducing strain,and S.cerevisiae acted as the helper strain.Comparative transcriptomic analysis was performed on an engineered S.cerevisiae(VTC2)and a wild-type S.cerevisiae BY4741.The results showed that the up-regulated genes in VTC2,compared with BY4741,were mainly involved in glycolysis,TCA cycle,purine metabolism,and biosynthesis of amino acids,B vitamins,and antioxidant proteases,all of which play important roles in pro-moting the growth of K.vulgare.Furthermore,Vitamin C produced by VTC2 could further relieve the oxidative stress in the environment to increase the production of 2-KLG.Therefore,VTC2 would be of great advantage in working with K.vulgare.Thus,the synthetic microbial consortium"VTC2-K.vulgare"was constructed based on transcriptomics analyses,and the accumulation of 2-KLG was increased by 1.49-fold compared with that of mono-cultured K.vulgare,reaching 13.2±0.52 g/L.In addition,the increased production of 2-KLG was accompanied by the up-regulated activities of superoxide dismutase and catalase in the medium and the up-regulated oxidative stress-related genes(sod,cat and gpd)in K.vulgare.The results indicated that the oxida-tive stress in the synthetic microbial consortium was efficiently reduced.Thus,systems analysis confirmed a favorable symbiotic relationship between microorganisms,providing guidance for further engineering synthetic consortia.

关键词： 2-Keto-L-gulonic acid S.cerevisiae K.vulgare Synthetic microbial consortia Transcriptomic analysis Synthetic biology

Enhancement and mapping of tolerance to salt stress and 5-fluorocytosine in synthetic yeast strains via SCRaMbLE

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Synthetic and Systems Biotechnology 2022年第3期7卷 869-877页

作者： Jianping Kang Jieyi Li Zhou Guo Sijie Zhou Shuxin Su Wenhai Xiao Yi Wu yingjin yuan Frontier Science Center for Synthetic Biology(Ministry of Education) Tianjin UniversityTianjin300072China Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin300072China

Varied environmental stress can affect cell growth and activity of the cellular catalyst.Traditional path of adaptive evolution generally takes a long time to achieve a tolerance phenotype,meanwhile,it is a challenge to dissect the underlying genetic mechanism.Here,using SCRaMbLE,a genome scale tool to generate random structural variations,a total of 222 evolved yeast strains with enhanced environmental tolerances were obtained in haploid or diploid yeasts containing six synthetic chromosomes.Whole genome sequencing of the evolved strains revealed that these strains generated different structural variants.Notably,by phenotypic-genotypic analysis of the SCRaMbLEd strains,we find that a deletion of gene YFR009W(GCN20)can improve salt tolerance of Saccharomyces cerevisiae,and a deletion of gene YER056C can improve 5-flucytosine tolerance of Saccharomyces cerevisiae.This study shows applications of SCRaMbLE to accelerate phenotypic evolution for varied environmental stress and to explore relationships between structural variations and evolved phenotypes.

关键词： SCRaMbLE Saccharomyces cerevisiae Structural variation GCN20 Tolerance

Simulating androgen receptor selection in designer yeast

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Synthetic and Systems Biotechnology 2022年第4期7卷 1108-1116页

作者： Haoran Zhang Lu Zhang Yipeng Xu Shaoyong Chen Zhenyi Ma Mingdong Yao Fangyin Li Bo Li yingjin yuan Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) Tianjin UniversityTianjin300072PR China School of Chemical Engineering and Technology Tianjin UniversityTianjin300072PR China Department of Urology The Cancer Hospital of the University of Chinese Academy of Sciences(Zhejiang Cancer Hospital)HangzhouChina Hematology-Oncology Division Department of MedicineBeth Israel Deaconess Medical Center and Harvard Medical SchoolBostonMA02215USA School of Basic Medical Sciences Tianjin Medical UniversityTianjin300070PR China

Androgen receptor(AR)mutation is closely associated with prostate cancer(PCa)and is one of the mechanisms of resistance to PCa therapies such as AR antagonists.Although sequencing technologies like next-generation sequencing(NGS)contributes to the high-throughput and precise detection of AR mutations carried by PCa patients,the lack of interpretations of these clinical genetic variants has still been a roadblock for PCa-targeted precision medicine.Here,we established a designer yeast reporter assay to simulate natural androgen receptor(AR)selection using AR antagonists.Yeast HIS3 gene transactivation was associated with the ligand-induced recruitment of steroid receptor coactivator-1(SRC-1)by AR mutants,where yeast growth in histidine-free medium was determined as the outcome.This assay is applicable to determine a wide range of clinical AR mutants including those with loss of function relating to androgen insensitivity syndrome(AIS),and those associated with PCa conferring resistance to AR antagonists such as enzalutamide(ENZ),bicalutamide(BIC),and cyproterone acetate(CPA).One clinical AR mutant previously reported to confer ENZ-resistance,F877L,was found to confer partial resistance to CPA as well using designer yeast.Our simple and efficient assay can enable precise one-pot screening of AR mutants,providing a reference for tailored medicine.

关键词： Androgen receptor mutations Androgen receptor antagonists Prostate cancer Saccharomyces cerevisiae

Enabling technology and core theory of synthetic biology

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Science China(Life Sciences) 2023年第8期66卷 1742-1785页

作者： Xian-En Zhang Chenli Liu Junbiao Dai yingjin yuan Caixia Gao Yan Feng Bian Wu Ping Wei Chun You Xiaowo Wang Tong Si Faculty of Synthetic Biology Shenzhen Institute of Advanced TechnologyShenzhen 518055China Institute of Synthetic Biology Shenzhen Institute of Advanced TechnologyChinese Academy of SciencesShenzhen 518055China National Laboratory of Biomacromolecules Institute of BiophysicsChinese Academy of SciencesBeijing 100101China Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin 300072China State Key Laboratory of Plant Cell and Chromosome Engineering Institute of Genetics and Developmental BiologyChinese Academy of SciencesBeijing 100101China State Key Laboratory of Microbial Metabolism Shanghai Jiao Tong UniversityShanghai 200240China State Key Laboratory of Microbial Resources Institute of MicrobiologyChinese Academy of SciencesBeijing 100101China Tianjin Institute of Industrial Biotechnology Chinese Academy of SciencesTianjin 300308China Ministry of Education Key Laboratory of Bioinformatics Center for Synthetic and Systems BiologyBioinformatics DivisionBeijing National Research Center for Information Science and TechnologyDepartment of AutomationTsinghua UniversityBeijing 100084China

Synthetic biology provides a new paradigm for life science research(“build to learn”)and opens the future journey of biotechnology(“build to use”).Here,we discuss advances of various principles and technologies in the mainstream of the enabling technology of synthetic biology,including synthesis and assembly of a genome,DNA storage,gene editing,molecular evolution and de novo design of function proteins,cell and gene circuit engineering,cell-free synthetic biology,artificial intelligence(AI)-aided synthetic biology,as well as biofoundries.We also introduce the concept of quantitative synthetic biology,which is guiding synthetic biology towards increased accuracy and predictability or the real rational design.We conclude that synthetic biology will establish its disciplinary system with the iterative development of enabling technologies and the maturity of the core theory.

关键词： synthetic biology quantitative synthetic biology genome synthesis and assembly DNA storage molecular evolution de novo design computer-aided design cell engineering gene circuit chassis cell artificial intelligent(AI) biofoundry

Collective Molecular Machines: Multidimensionality and Reconfigurability

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Nano-Micro Letters 2024年第8期16卷 309-340页

作者： Bin Wang yuan Lu Department of Chemical Engineering Tsinghua UniversityBeijing 100084People’s Republic of China Key Laboratory of Industrial Biocatalysis Ministry of EducationTsinghua UniversityBeijing 100084People’s Republic of China

Molecular machines are key to cellular activity where they are involved in converting chemical and light energy into efficient mechanical work.During the last 60 years,designing molecular structures capable of generating unidirectional mechanical motion at the nanoscale has been the topic of intense research.Effective progress has been made,attributed to advances in various fields such as supramolecular chemistry,biology and nanotechnology,and informatics.However,individual molecular machines are only capable of producing nanometer work and generally have only a single functionality.In order to address these problems,collective behaviors realized by integrating several or more of these individual mechanical units in space and time have become a new paradigm.In this review,we comprehensively discuss recent developments in the collective behaviors of molecular machines.In particular,collective behavior is divided into two paradigms.One is the appropriate integration of molecular machines to efficiently amplify molecular motions and deformations to construct novel functional materials.The other is the construction of swarming modes at the supramolecular level to perform nanoscale or microscale operations.We discuss design strategies for both modes and focus on the modulation of features and properties.Subsequently,in order to address existing challenges,the idea of transferring experience gained in the field of micro/nano robotics is presented,offering prospects for future developments in the collective behavior of molecular machines.

关键词： Molecular machines Collective control Collective behaviors DNA Biomolecular motors