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Synthetic biology, an emerging research field, can promote biomanufacturing 4.0 by offering various efficient chassis. Engineering Bacillus subtilis, a prominent workhorse in industrial biotechnology, through synthetic biology approaches may be a disruptive innovation. Diverse chassis forms are envisaged to deal with challenges in metabolic complexity, metabolic burden, and systems robustness. Thus, advancements in chassis engineering, a synthetic biology strategy for genome‐reduced cell factories, cell‐free systems, and synthetic microbial consortia would be a driving force facilitating microbial production. In this review article, we summarized chassis engineering categories and applications for B. subtilis. Prospects and challenges for chassis engineering in B. subtilis were also discussed. © 2022 Society of Chemical Industry (SCI). [ABSTRACT FROM AUTHOR]

Background. Prostate cancer (PCa) is one of the most common malignancies in men. Increasing evidence has demonstrated that dysregulation of long noncoding RNAs (lncRNAs) is closely related to carcinogenesis and cancer progression. lncRNA NEAT1 has recently been identified as a carcinogenic regulator of multiple cancers; however, the role of NEAT1 on PCa is still poorly understood. Methods. Kaplan–Meier was conducted to determine the overall survival rate in PCa patients with aberrant NEAT1 levels. qRT-PCR analysis was performed to detect expressions of NEAT1 and miR-766-5p in tissues and cells. In addition, CCK-8, colony formation, flow cytometry analysis, wound healing, and transwell assay were conducted to determine cell proliferation, cell arrest, apoptosis, migration, and invasion. The western blot assay was utilized to determine E2F3 and cell growth-related proteins. The relationship between NEAT1 and miR-766-5p or miR-766-5p and E2F3 was verified by correlation analysis and dual-luciferase reporter assay. Results. Here, we find that NEAT1 is overexpressed in PCa tissues and cell lines. Besides, silencing of NEAT1 inhibits cell proliferation, invasion, and migration and promotes cell apoptosis and cell cycle arrest. Further mechanistic studies find that NEAT1 sponges miR-766-5p, and miRNA-766-5p is negatively correlated with the expression of NEAT1. In addition, the functional experiment shows that upregulation of miRNA-766-5p inhibits PCa proliferation, migration, and invasion. Furthermore, E2F transcription factor 3 (E2F3) is testified to be the downstream target gene of miRNA-766-5p. Finally, the rescue experiment revealed that miRNA-766-5p inhibition largely restores NEAT1 downregulation-mediated function on PCa progression, while E2F3 knockdown partly removes the effects of miRNA-766-5p inhibitor. Conclusions. In conclusion, NEAT1 facilitates PCa progression by targeting the miRNA-766-5p/E2F3 axis. [ABSTRACT FROM AUTHOR]

A new protocol for metabolomic studies was developed by combining liquid chromatography-tandem mass spectrometry and isotope dilution mass spectrometry with universal13C labeled internal standards fromEscherichia Coli. The multiple reaction monitoring mode of mass spectrometry was used for quantification. Forty-five water-soluble intracellular metabolites, including 20 amino acids, 16 organic acids (primarily from the tricarboxylic acid cycle), and 9 cofactors, were measured and 34 of them were successfully quantified using the13C-labeled internal standards. The limit of detection, limit of quantification, precision, and linearity of the methods were evaluated. The methods were applied to the quantitative analysis of intracellular metabolites extracted from wild-type and ethanol-adapted strains ofClostridium thermocellumcultivated with and without ethanol stress, and all 34 metabolites including all 9 cofactors were successfully quantified. Further multivariate data analyses of the metabolic differences between wild-type and ethanol-adapted strains were performed on the quantitative data, which can help elucidate the metabolic mechanism behind ethanol adaptation inC. thermocellum. Supplemental materials are available for this article. Go to the publisher's online edition ofAnalytical Lettersto view the supplemental file. [ABSTRACT FROM AUTHOR]

Clostridium thermocellum is a major candidate for bioethanol production via consolidated bioprocessing. However, the low ethanol tolerance of the organism dramatically impedes its usage in industry. To explore the mechanism of ethanol tolerance in this microorganism, systematic metabolomics was adopted to analyse the metabolic phenotypes of a C. thermocellum wild-type (WT) strain and an ethanol-tolerant strain cultivated without (ET0) or with (ET3) 3% (v/v) exogenous ethanol. Metabolomics analysis elucidated that the levels of numerous metabolites in different pathways were changed for the metabolic adaption of ethanol-tolerant C. thermocellum. The most interesting phenomenon was that cellodextrin was significantly more accumulated in the ethanol-tolerant strain compared with the WT strain, although cellobiose was completely consumed in both the ethanol-tolerant and wild-type strains. These results suggest that the cellodextrin synthesis was active, which might be a potential mechanism for stress resistance. Moreover, the overflow of many intermediate metabolites, which indicates the metabolic imbalance, in the ET0 cultivation was more significant than in the WT and ET3 cultivations. This indicates that the metabolic balance of the ethanol-tolerant strain was adapted better to the condition of ethanol stress. This study provides additional insight into the mechanism of ethanol tolerance and is valuable for further metabolic engineering aimed at higher bioethanol production. [ABSTRACT FROM AUTHOR]