生物设计研究(英文)最新文献

Terpenoids of substantial industrial interest are mainly obtained through direct extraction from plant sources. Recently, microbial cell factories or in vitro enzymatic biosystems have emerged as promising alternatives for terpenoid production. Here, we report a route for the synthesis of α-farnesene based on an in vitro enzyme cascade reaction using methanol as an inexpensive and renewable C1 substrate. Thirteen biocatalytic reactions divided into 2 modules were optimized and coupled to achieve methanol-to-α-farnesene conversion via integration with natural thylakoid membranes as a green energy engine. This in vitro enzymatic biosystem driven by light enabled the production of 1.43 and 2.40 mg liter^-1 α-farnesene using methanol and the intermediate glycolaldehyde as substrates, respectively. This work could provide a promising strategy for developing light-powered in vitro biosynthetic platforms to produce more natural compounds synthesized from C1 substrates.

Recently, there has been increasing interest in the use of bacteria for cancer therapy due to their ability to selectively target tumor sites and inhibit tumor growth. However, the complexity of the interaction between bacteria and tumor cells evokes unpredictable therapeutic risk, which induces inflammation, stimulates the up-regulation of cyclooxygenase II (COX-2) protein, and stimulates downstream antiapoptotic gene expression in the tumor microenvironment to reduce the antitumor efficacy of chemotherapy and immunotherapy. In this study, we encapsulated celecoxib (CXB), a specific COX-2 inhibitor, in liposomes anchored to the surface of Escherichia coli Nissle 1917 (ECN) through electrostatic absorption (C@ECN) to suppress ECN-induced COX-2 up-regulation and enhance the synergistic antitumor effect of doxorubicin (DOX). C@ECN improved the antitumor effect of DOX by restraining COX-2 expression. In addition, local T lymphocyte infiltration was induced by the ECN to enhance immunotherapy efficacy in the tumor microenvironment. Considering the biosafety of C@ECN, a hypoxia-induced lysis circuit, pGEX-Pvhb-Lysis, was introduced into the ECN to limit the number of ECNs in vivo. Our results indicate that this system has the potential to enhance the synergistic effect of ECN with chemical drugs to inhibit tumor progression in medical oncology.

In the biotechnological industry, multicopy gene integration represents an effective strategy to maintain a high-level production of recombinant proteins and to assemble multigene biochemical pathways. In this study, we developed copper-induced in vivo gene amplification in budding yeast for multicopy gene expressions. To make copper as an effective selection pressure, we first constructed a copper-sensitive yeast strain by deleting the CUP1 gene encoding a small metallothionein-like protein for copper resistance. Subsequently, the reporter gene fused with a proline-glutamate-serine-threonine-destabilized CUP1 was integrated at the δ sites of retrotransposon (Ty) elements to counter the copper toxicity at 100 μM Cu²⁺. We further demonstrated the feasibility of modulating chromosomal rearrangements for increased protein expression under higher copper concentrations. In addition, we also demonstrated a simplified design of integrating the expression cassette at the CUP1 locus to achieve tandem duplication under high concentrations of copper. Taken together, we envision that this method of copper-induced in vivo gene amplification would serve as a robust and useful method for protein overproduction and metabolic engineering applications in budding yeast.

Plants are complex systems hierarchically organized and composed of various cell types. To understand the molecular underpinnings of complex plant systems, single-cell RNA sequencing (scRNA-seq) has emerged as a powerful tool for revealing high resolution of gene expression patterns at the cellular level and investigating the cell-type heterogeneity. Furthermore, scRNA-seq analysis of plant biosystems has great potential for generating new knowledge to inform plant biosystems design and synthetic biology, which aims to modify plants genetically/epigenetically through genome editing, engineering, or re-writing based on rational design for increasing crop yield and quality, promoting the bioeconomy and enhancing environmental sustainability. In particular, data from scRNA-seq studies can be utilized to facilitate the development of high-precision Build-Design-Test-Learn capabilities for maximizing the targeted performance of engineered plant biosystems while minimizing unintended side effects. To date, scRNA-seq has been demonstrated in a limited number of plant species, including model plants (e.g., Arabidopsis thaliana), agricultural crops (e.g., Oryza sativa), and bioenergy crops (e.g., Populus spp.). It is expected that future technical advancements will reduce the cost of scRNA-seq and consequently accelerate the application of this emerging technology in plants. In this review, we summarize current technical advancements in plant scRNA-seq, including sample preparation, sequencing, and data analysis, to provide guidance on how to choose the appropriate scRNA-seq methods for different types of plant samples. We then highlight various applications of scRNA-seq in both plant systems biology and plant synthetic biology research. Finally, we discuss the challenges and opportunities for the application of scRNA-seq in plants.

The golden age has passed for antibiotic discovery, and while some antibiotics are currently in various phases of clinical trials in the United States, many pharmaceutical companies have abandoned antibiotic research. With the need for antibiotics, we should expand our horizon for therapeutic mining and can look toward understudied sources such as ice cores. Ice cores contain microorganisms and genetic material that have been frozen in time for thousands of years. The antibiotics used by these organisms are encoded in their genomes, which can be unlocked, identified, and characterized with modern advances in molecular biology, genetic sequencing, various computational approaches, and established natural product discovery pipelines. While synthetic biology can be used in natural product discovery approaches, synthetic biology and bioengineering efforts can also be leveraged in the selection and biodesign of increased compound yields, potency, and stability. Here, we provide the perspective that ice cores can be a source of novel antibiotic compounds and that the tools of synthetic biology can be used to design better antimicrobials.

The third-generation (3G) biorefinery aims to use microbial cell factories or enzymatic systems to synthesize value-added chemicals from one-carbon (C1) sources, such as CO₂, formate, and methanol, fueled by renewable energies like light and electricity. This promising technology represents an important step toward sustainable development, which can help address some of the most pressing environmental challenges faced by modern society. However, to establish processes competitive with the petroleum industry, it is crucial to determine the most viable pathways for C1 utilization and productivity and yield of the target products. In this review, we discuss the progresses that have been made in constructing artificial biological systems for 3G biorefineries in the last 10 years. Specifically, we highlight the representative works on the engineering of artificial autotrophic microorganisms, tandem enzymatic systems, and chemo-bio hybrid systems for C1 utilization. We also prospect the revolutionary impact of these developments on biotechnology. By harnessing the power of 3G biorefinery, scientists are establishing a new frontier that could potentially revolutionize our approach to industrial production and pave the way for a more sustainable future.

Probiotics are the treasure of the microbiology fields. They have been widely used in the food industry, clinical treatment, and other fields. The equivocal health-promoting effects and the unknown action mechanism were the largest obstacles for further probiotic's developed applications. In recent years, various genome editing techniques have been developed and applied to explore the mechanisms and functional modifications of probiotics. As important genome editing tools, CRISPR-Cas systems that have opened new improvements in genome editing dedicated to probiotics. The high efficiency, flexibility, and specificity are the advantages of using CRISPR-Cas systems. Here, we summarize the classification and distribution of CRISPR-Cas systems in probiotics, as well as the editing tools developed on the basis of them. Then, we discuss the genome editing of probiotics based on CRISPR-Cas systems and the applications of the engineered probiotics through CRISPR-Cas systems. Finally, we proposed a design route for CRISPR systems that related to the genetically engineered probiotics.

Broad-host-range synthetic biology is an emerging frontier that aims to expand our current engineerable domain of microbial hosts for biodesign applications. As more novel species are brought to "model status," synthetic biologists are discovering that identically engineered genetic circuits can exhibit different performances depending on the organism it operates within, an observation referred to as the "chassis effect." It remains a major challenge to uncover which genome-encoded and biological determinants will underpin chassis effects that govern the performance of engineered genetic devices. In this study, we compared model and novel bacterial hosts to ask whether phylogenomic relatedness or similarity in host physiology is a better predictor of genetic circuit performance. This was accomplished using a comparative framework based on multivariate statistical approaches to systematically demonstrate the chassis effect and characterize the performance dynamics of a genetic inverter circuit operating within 6 Gammaproteobacteria. Our results solidify the notion that genetic devices are strongly impacted by the host context. Furthermore, we formally determined that hosts exhibiting more similar metrics of growth and molecular physiology also exhibit more similar performance of the genetic inverter, indicating that specific bacterial physiology underpins measurable chassis effects. The result of this study contributes to the field of broad-host-range synthetic biology by lending increased predictive power to the implementation of genetic devices in less-established microbial hosts.

High-precision bioengineering and synthetic biology require fine-tuning gene expression at both transcriptional and posttranscriptional levels. Gene transcription is tightly regulated by promoters and terminators. Promoters determine the timing, tissues and cells, and levels of the expression of genes. Terminators mediate transcription termination of genes and affect mRNA levels posttranscriptionally, e.g., the 3'-end processing, stability, translation efficiency, and nuclear to cytoplasmic export of mRNAs. The promoter and terminator combination affects gene expression. In the present article, we review the function and features of plant core promoters, proximal and distal promoters, and terminators, and their effects on and benchmarking strategies for regulating gene expression.