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Abstract Base editors, which enable targeted locus nucleotide conversion in genomic DNA without double‐stranded breaks, have been engineered as powerful tools for biotechnological and clinical applications. However, the application of base editors is limited by their off‐target effects. Continuously expressed deaminases used for gene editing may lead to unwanted base alterations at unpredictable genomic locations. In the present study, blue‐light‐activated base editors (BLBEs) are engineered based on the distinct photoswitches magnets that can switch from a monomer to dimerization state in response to blue light. By fusing the N‐ and C‐termini of split DNA deaminases with photoswitches Magnets, efficient A‐to‐G and C‐to‐T base editing is achieved in response to blue light in prokaryotic and eukaryotic cells. Furthermore, the results showed that BLBEs can realize precise blue light‐induced gene editing across broad genomic loci with low off‐target activity at the DNA‐ and RNA‐level. Collectively, these findings suggest that the optogenetic utilization of base editing and optical base editors may provide powerful tools to promote the development of optogenetic genome engineering.

Abstract Background Wood-feeding termite, Coptotermes formosanus Shiraki, represents a highly efficient system for biomass deconstruction and utilization. However, the detailed mechanisms of lignin modification and carbohydrate degradation in this system are still largely elusive. Results In order to reveal the inherent mechanisms for efficient biomass degradation, four different organs (salivary glands, foregut, midgut, and hindgut) within a complete digestive system of a lower termite, C. formosanus, were dissected and collected. Comparative transcriptomics was carried out to analyze these organs using high-throughput RNA sequencing. A total of 71,117 unigenes were successfully assembled, and the comparative transcriptome analyses revealed significant differential distributions of GH (glycosyl hydrolase) genes and auxiliary redox enzyme genes in different digestive organs. Among the GH genes in the salivary glands, the most abundant were GH9, GH22, and GH1 genes. The corresponding enzymes may have secreted into the foregut and midgut to initiate the hydrolysis of biomass and to achieve a lignin-carbohydrate co-deconstruction system. As the most diverse GH families, GH7 and GH5 were primarily identified from the symbiotic protists in the hindgut. These enzymes could play a synergistic role with the endogenous enzymes from the host termite for biomass degradation. Moreover, twelve out of fourteen genes coding auxiliary redox enzymes from the host termite origin were induced by the feeding of lignin-rich diets. This indicated that these genes may be involved in lignin component deconstruction with its redox network during biomass pretreatment. Conclusion These findings demonstrate that the termite digestive system synergized the hydrolysis and redox reactions in a programmatic process, through different parts of its gut system, to achieve a maximized utilization of carbohydrates. The detailed unique mechanisms identified from the termite digestive system may provide new insights for advanced design of future biorefinery.

Abstract Neuronal cells express considerable plasticity responding to environmental cues, in part, through subcellular mRNA regulation. Here we report on the extensive changes in distribution of mRNAs in the cell body and axon compartments of peripheral sensory neurons and the 3′ untranslated region (3′UTR) landscapes after unilateral sciatic nerve entrapment (SNE) injury in rats. Neuronal cells dissociated from SNE-injured and contralateral L4 and L5 dorsal root ganglia were cultured in a compartmentalized system. Axonal and cell body RNA samples were separately subjected to high throughput RNA sequencing (RNA-Seq). The injured axons exhibited enrichment of mRNAs related to protein synthesis and nerve regeneration. Lengthening of 3′UTRs was more prevalent in the injured axons, including the newly discovered alternative cleavage and polyadenylation of NaV1.8 mRNA. Alternative polyadenylation was largely independent from the relative abundance of axonal mRNAs; but they were highly clustered in functional pathways related to RNA granule formation in the injured axons. These RNA-Seq data analyses indicate that peripheral nerve injury may result in highly selective mRNA enrichment in the affected axons with 3′UTR alterations potentially contributing to the mechanism of neuropathic pain.

Abstract Bacterial protein secretion represents a significant challenge in biotechnology, which is essential for the cost‐effective production of therapeutics, enzymes, and other functional proteins. Here, it is demonstrated that proteomics‐guided engineering of transcription, translation, secretion, and folding of ligninolytic laccase balances the process, minimizes the toxicity, and enables efficient heterologous secretion with a total protein yield of 13.7 g L−1. The secretory laccase complements the biochemical limits on lignin depolymerization well in Rhodococcus opacus PD630. Further proteomics analysis reveals the mechanisms for the oleaginous phenotype of R. opacus PD630, where a distinct multiunit fatty acid synthase I drives the carbon partition to storage lipid. The discovery guides the design of efficient lipid conversion from lignin and carbohydrate. The proteomics‐guided integration of laccase‐secretion and lipid production modules enables a high titer in converting lignin‐enriched biorefinery waste to lipid. The fundamental mechanisms, engineering components, and design principle can empower transformative platforms for biomanufacturing and biorefining.

Terpenes are the major secondary metabolites produced by plants, and have diverse industrial applications as pharmaceuticals, fragrance, solvents, and biofuels. Cyanobacteria are equipped with efficient carbon fixation mechanism, and are ideal cell factories to produce various fuel and chemical products. Past efforts to produce terpenes in photosynthetic organisms have gained only limited success. Here we engineered the cyanobacterium Synechococcus elongatus PCC 7942 to efficiently produce limonene through modeling guided study. Computational modeling of limonene flux in response to photosynthetic output has revealed the downstream terpene synthase as a key metabolic flux-controlling node in the MEP (2-C-methyl-d-erythritol 4-phosphate) pathway-derived terpene biosynthesis. By enhancing the downstream limonene carbon sink, we achieved over 100-fold increase in limonene productivity, in contrast to the marginal increase achieved through stepwise metabolic engineering. The establishment of a strong limonene flux revealed potential synergy between photosynthate output and terpene biosynthesis, leading to enhanced carbon flux into the MEP pathway. Moreover, we show that enhanced limonene flux would lead to NADPH accumulation, and slow down photosynthesis electron flow. Fine-tuning ATP/NADPH toward terpene biosynthesis could be a key parameter to adapt photosynthesis to support biofuel/bioproduct production in cyanobacteria. [ABSTRACT FROM AUTHOR]

Background Current knowledge about the molecular properties and prognostic markers of upper tract urothelial carcinoma (UTUC) is sparse and often based on bladder urothelial carcinoma (UC), which is thought to share common risk factors with UTUC. However, studies have suggested that differences exist regarding tumor behavior and molecular biology of these cancers, comprehensive investigations are needed to guide the clinical management of UTUC. In recent years, massively parallel sequencing has allowed insights into the biology of many cancers, and molecular prognostic markers based on this approach are rapidly emerging. The goal of this study was to characterize the gene expression patterns of UTUC using massively parallel sequencing, and identify potential molecular markers for prognosis in patients with UTUC. Methods We compared the genome-wide mRNA expression profile of cancer and matched normal tissues from 10 patients with UTUC to identify significantly deregulated genes. We also examined the protein levels of prognostic marker candidates in 103 patients with UTUC, and tested the association of these markers with overall survival using Kaplan-Meier model and Cox regression. Results Functional enrichment of significantly deregulated genes revealed that expression patterns of UTUC were characterized by disorders of cell proliferation and metabolism. And we also compared the expression profile of UTUC with that of bladder UC. Our results highlighted both shared (e.g. disorders of cell cycling and growth signal transduction) and tumor-specific (e.g. abnormal metabolism in UTUC and disruptions of adhesion pathways in bladder UC) features of these two cancers. Importantly, we identified that low protein expression of ALDH2 while high CCNE1 and SMAD3 were significantly associated with increased depth (*P <0.05) and lower overall survival (***P <0.0001) in an independent set of 103 patients. Multivariate Cox regression revealed that all these three genes were independent prognostic indicators in patients with UTUC (***P <0.001). Conclusions In conclusion, our study characterized the comprehensive expression profile of UTUC and highlighted both commons and differences in expression patterns between UTUC and bladder UC. And we, for the first time, revealed that ALDH2, CCNE1 and SMAD3 are associated with prognosis in patients with UTUC. [ABSTRACT FROM AUTHOR]

The article focuses on a study which identified the representative genes of the central nervous system (CNS) of the locust Locusta migratoria manilensis using deep sequencing. The study identified 15,895 unigenes from the Locusta migratoria manilensis, including 644 nervous system relevant unigenes. It noted that the locust CNS matches those of the Tribolium castaneum (Coleoptera: Tenebrionidae) sequences.

Polyadenylated mature mRNAs are the focus of standard transcriptome analyses. However, the profiling of nascent transcripts, which often include nonpolyadenylated RNAs, can unveil novel insights into transcriptional regulation. Here, we separately sequenced total RNAs (Total RNAseq) and mRNAs (mRNAseq) from the same HIV-1-infected human CD4+ T cells. We found that many nonpolyadenylated RNAs were differentially expressed upon HIV-1 infection, and we identified 8 times more differentially expressed genes at 12 h postinfection by Total RNAseq than by mRNAseq. These expression changes were also evident by concurrent changes in introns and were recapitulated by later mRNA changes, revealing an unexpectedly significant delay between transcriptional initiation and mature mRNA production early after HIV-1 infection. We computationally derived and validated the underlying regulatory programs, and we predicted drugs capable of reversing these HIV-1-induced expression changes followed by experimental confirmation. Our results show that combined total and mRNA transcriptome analysis is essential for fully capturing the early host response to virus infection and provide a framework for identifying candidate drugs for host-directed therapy against HIV/AIDS. [ABSTRACT FROM AUTHOR]