Science Advances
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2019
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IF 12.804
Yongchun Pan,Jingjing Yang,Xiaowei Luan,Xinli Liu,Xueqing Li,Jian Yang,Ting Huang,Lu Sun,Yuzhen Wang,Youhui Lin,Yujun Song
As an RNA-guided nuclease, CRISPR-Cas9 offers facile and promising solutions to mediate genome modification with respect to versatility and high precision. However, spatiotemporal manipulation of CRISPR-Cas9 delivery remains a daunting challenge for robust effectuation of gene editing both in vitro and in vivo. Here, we designed a near-infrared (NIR) light–responsive nanocarrier of CRISPR-Cas9 for cancer therapeutics based on upconversion nanoparticles (UCNPs). The UCNPs served as “nanotransducers” that can convert NIR light (980 nm) into local ultraviolet light for the cleavage of photosensitive molecules, thereby resulting in on-demand release of CRISPR-Cas9. In addition, by preparing a single guide RNA targeting a tumor gene (polo-like kinase-1), our strategies have successfully inhibited the proliferation of tumor cell via NIR light–activated gene editing both in vitro and in vivo. Overall, this exogenously controlled method presents enormous potential for targeted gene editing in deep tissues and treatment of a myriad of diseases.
研究领域:
遗传学
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纳米技术
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癌症研究
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分子生物学
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高影响因子
ANALYTICA CHIMICA ACTA
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2026
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IF 6.1
Wen Xu,Yutian Cheng,Kexin Sun,Yue Wu,Yiming Xu,Jiang Ye,Pengfei Li,Haizhen Wu
Background Kanamycin's persistent contamination in agricultural products poses significant human health risks due to its nephrotoxicity and bioaccumulation via the food chain. Existing methods for on-site kanamycin monitoring lack sufficient sensitivity and portability, limiting their utility in field settings. The urgent need for rapid detection technologies remains unmet. This work addresses the critical gap in developing a field-deployable platform for ultrasensitive kanamycin residue screening. Results We engineered a fluorescence biosensor integrating DNAzyme-assisted Cas9 nickase-based amplification reaction (Cas9nAR). Kanamycin binding induces aptamer conformational changes, triggering Cas9nAR-driven cascade amplification that continuously generates DNAzymes. These cleave reporter probes to enable quantitative detection. The system achieved a broad linear range (1 nM – 5 μM) with an ultralow detection limit (0.3 nM), surpassing conventional methods. It exhibited high specificity against interfering antibiotics and delivered consistent recoveries (97% to 103%) in spiked water and milk samples. Following pretreatment, analysis can be completed within 120 min, validating its operational simplicity and robustness for complex matrices. Significance and novelty This work reports the first CRISPR-Cas9 nickase/DNAzyme cascade amplification platform for small-molecule detection, establishing a new paradigm that integrates programmable nucleic acid amplification with catalytic signal turnover. By using Cas9nAR-generated ssDNA as an in situ template for autonomous DNAzyme assembly, the biosensor achieves ultrasensitive, and homogeneous detection of kanamycin-addressing a critical gap in field-deployable antibiotic residue monitoring. The modular design offers a generalizable strategy for translating non-nucleic acid recognition events into amplified fluorescent outputs, with broad implications for point-of-need diagnostics in food safety and environmental analysis.
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Journal of Fungi
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2023
·Vol. 9
·期 8
·863
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IF 4.7
Yinggu Wu,Xinyu Li,Li Dong,Tong Liu,Zhengbin Tang,Runmao Lin,Justice Norvienyeku,Mengyu Xing
The litchi downy blight disease of litchi caused byPeronophythora litchiiaccounts for severe losses in the field and during storage. While ample quantitative studies have shown that 6-pentyl-2H-pyran-2-one (6PP) possesses antifungal activities against multiple plant pathogenic fungi, the regulatory mechanisms of 6PP-mediated inhibition of fungal pathogenesis and growth are still unknown. Here, we investigated the potential molecular targets of 6PP in the phytopathogenic oomycetesP. litchiithrough integrated deployment of RNA-sequencing, functional genetics, and biochemical techniques to investigate the regulatory effects of 6PP againstP. litchii. Previously we demonstrated that 6PP exerted significant oomyticidal activities. Also, comparative transcriptomic evaluation ofP. litchiistrains treated with 6PP Revealed significant up-regulations in the expression profile of TOR pathway-related genes, includingPlCytochrome Cand the transcription factorsPlYY1. We also noticed that 6PP treatment down-regulated putative negative regulatory genes of the TOR pathway, includingPlSpm1andPlrhoH12inP. litchii. Protein-ligand binding analyses revealed stable affinities between PlYY1, PlCytochrome C, PlSpm1, PlrhoH12 proteins, and the 6PP ligand. Phenotypic characterization ofPlYY1targeted gene deletion strains generated in this study using CRISPR/Cas9 and homologous recombination strategies significantly reduced the vegetative growth, sporangium, encystment, zoospore release, and pathogenicity ofP. litchii. These findings suggest that 6PP-mediated activation ofPlYY1expression positively regulates TOR-related responses and significantly influences vegetative growth and the virulence ofP. litchii. The current investigations revealed novel targets for 6PP and underscored the potential of deploying 6PP in developing management strategies for controlling the litchi downy blight pathogen.Keywords:P. litchii;6-pentyl-2H-pyran-2-one (6PP);TOR pathway;CRISPR/Cas 9;PlYY1
研究领域:
植物病理学
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分子生物学
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真菌学
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BIOMEDICINE & PHARMACOTHERAPY
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2022
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IF 7.419
Qingxia Zuo,Wanqing Xu,Yanbin Wan,Dongyan Feng,Changsheng He,Cailing Lin,Dongchao Huang,Feng Chen,Liya Han,Qi Sun,Dong Chen,Hongli Du,Lizhen Huang
CRISPR/Cas9 has allowed development of better and easier-to-use ADME models than traditional methods by complete knockout or knock-in of genes. However, gene editing in HepaRG cells remains challenging because long-term monoclonal cultivation may alter their differentiation capacity to a large extent. Here, CRISPR/Cas9 was used to generate a CYP3A4-T2A-luciferase knock-in HepaRG subclone by Cas9-mediated homologous recombination and monoclonal cultivation. The knock-in HepaRG-#9 subclone retained a similar differentiation potential to wildtype HepaRG cells (HepaRG-WT). To further improve differentiation and expand the applications of knock-in HepaRG cells, two optimized differentiation procedures were evaluated by comparison with the standard differentiation procedure using the knock-in HepaRG-#9 subclone and HepaRG-WT. The results indicated that addition of forskolin (an adenylate cyclase activator) and SB431542 (a TGF-β pathway inhibitor) to the first optimized differentiation procedure led to better differentiation consequence in terms of not only the initiation time for differentiation and morphological characterization, but also the mRNA levels of hepatocyte-specific genes. These data may contribute to more extensive applications of genetically modified HepaRG cells in ADME studies.
研究领域:
药理学
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分子生物学
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基因工程
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PubMed
高影响因子
Cell Reports Medicine
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2026
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IF 14
Chen Zhang,Tianyu Li,Huoye Hao,Gang Wang,Yuzhe Zhang,Yue Huang,Lili Chen,Jin Wang,Xiang-Jiao Yang,Xiuli Bi
The primary pathogenic mechanism of inflammatory bowel disease (IBD) involves elevated levels of reactive oxide species (ROS) in the gut, leading to oxidative stress and damage to the intestinal barrier function. We engineered a non-pathogenic bacterial strain, Escherichia coli Nissle 1917 (EcN), for oral CRISPR-Cas9 delivery to edit NOX2 (encoding the gp91 phox subunit of NADPH oxidase 2), thereby alleviating IBD symptoms by reducing ROS. EcN expressing the Cas9/sgRNA ribonucleoprotein (RNP) was encapsulated in a hydrogel (composed of hyaluronic acid, chitosan, and MgCl 2 ), which protected EcN-RNP from degradation and increased survival from 0.07% to 12%. EcN-RNP hydrogel reduced NOX2 expression by 47% and ROS by 88% in lipopolysaccharide-induced RAW264.7 cells. Moreover, oral delivery of EcN-RNP hydrogel mitigated inflammation in dextran sodium sulfate-induced colitis mouse models. Mechanistically, the hydrogel could activate the NRF2-HO1/GPX4 pathway by inhibiting NOX2 expression, enhancing oxidative defense, and promoting glutathione accumulation. Thus, the EcN-RNP hydrogel offers a promising therapeutic strategy for IBD.
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Lipids in Health and Disease
·
2021
·Vol. 20
·期 1
·1-11
·
IF 3.876
Zha Yiwen,Lu Yaoyao,Zhang Ting,Yan Kunning,Zhuang Wenwen,Liang Jingyan,Cheng Yong,Wang Yingge
Background High levels of apolipoprotein C3 (APOC3) can lead to hypertriglyceridemia, which increases the risk of cardiovascular disease. We aim to create APOC3-knockout (KO) rabbits and explore the effects of APOC3 deletion on the occurrence and development of atherosclerosis.Methods An sgRNA anchored to exon 2 of APOC3 was designed to edit embryo genomes using the CRISPR/Cas9 system. The founder rabbits were sequenced, and their lipid profile, inflammatory cytokines, and atherosclerotic plaques were analyzed.Results When given a normal chow (NC) diet, all APOC3-KO rabbits had 50% lower triglyceride (TG) levels than those of the matched age control group. Additionally, their plasma lipoprotein lipase increased. When fed a high-fat diet, APOC3 deficiency was observed to be more conducive to the maintenance of plasma TG, total cholesterol, and low-density lipoprotein cholesterol levels, and the inhibition of the inflammatory response and the protection against atherosclerosis in rabbits.Conclusion APOC3 deficiency can delay the formation of atherosclerosis-induced HFD in rabbits, indicating this is a novel therapeutic target to treat atherosclerosis.
研究领域:
分子生物学
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心血管疾病
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基因工程
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PubMed
高影响因子
Scientific Reports
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2017
·Vol. 7
·期 1
·1-8
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IF 4.259
Guo Dongsheng,Liu Haikun,Ruzi Aynisahan,Gao Ge,Nasir Abbas,Liu Yanli,Yang Fan,Wu Feima,Xu Guosheng,Li Yin-xiong
Congenital hyperinsulinism (CHI) is a rare genetic disorder characterized by excess insulin secretion, which results in hypoglycemia. Mutation of sulfonylurea receptor 1 (SUR1), encoded by the ABCC8 gene, is the main cause of CHI. Here, we captured the phenotype of excess insulin secretion through pancreatic differentiation of ABCC8 -deficient stem cells generated by the CRISPR/Cas9 system. ABCC8- deficient insulin-producing cells secreted higher insulin than their wild-type counterparts, and the excess insulin secretion was rescued by nifedipine, octreotide and nicorandil. Further, we tested the role of SUR1 in response to different potassium levels and found that dysfunction of SUR1 decreased the insulin secretion rate in low and high potassium environments. Hence, pancreatic differentiation of ABCC8- deficient cells recapitulated the CHI disease phenotype in vitro , which represents an attractive model to further elucidate the function of SUR1 and to develop and screen for novel therapeutic drugs.
研究领域:
遗传学
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干细胞研究
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分子生物学
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内分泌学
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DOI
PubMed
高影响因子
Nature Nanotechnology
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2025
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IF 34.9
Zhao Kai,Yan Yu,Jin Xiao-Kang,Pan Ting,Zhang Shi-Man,Yang Chi-Hui,Rao Zhi-Yong,Zhang Xian-Zheng
Chemoresistance and immunosuppression are common obstacles to the efficacy of chemo-immunotherapy in colorectal cancer (CRC) and are regulated by mitochondrial chaperone proteins. Here we show that the disruption of the tumour necrosis factor receptor-associated protein 1 ( TRAP1 ) gene, which encodes a mitochondrial chaperone in tumour cells, causes the translocation of cyclophilin D in tumour cells. This process results in the continuous opening of the mitochondrial permeability transition pore, which enhances chemotherapy-induced cell necrosis and promotes immune responses. On the basis of this discovery we developed an oral CRISPR–Cas9 delivery system based on zwitterionic and polysaccharide polymer-coated nanocomplexes that disrupts the TRAP1 gene in CRC. This system penetrates the intestinal mucus layer and undergoes epithelial transcytosis, accumulating in CRC tissues. It enhances chemotherapeutic efficacy by overcoming chemoresistance and activating the tumour immune microenvironment in orthotopic, chemoresistant and spontaneous CRC models, with remarkable synergistic antitumour effects. This oral CRISPR–Cas9 delivery system represents a promising therapeutic strategy for the clinical management of CRC.
研究领域:
肿瘤学
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分子生物学
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基因编辑
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纳米医学
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产品关联
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PubMed
高影响因子
PARASITOLOGY RESEARCH
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2019
·Vol. 118
·期 3
·935-944
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IF 2.067
Ye Shu,Xia Ningbo,Zhao Pengfei,Yang Jichao,Zhou Yanqin,Shen Bang,Zhao Junlong
Toxoplasma gondii is a ubiquitous parasitic protozoan infecting humans and a wide variety of animals. Fast-replicating tachyzoites during acute infection and slowly growing bradyzoites during chronic infection are the two basic forms of T. gondii in intermediate hosts. Interconversion between the two contributes to the transmission and pathogenesis of this parasite. Secretory micronemal proteins are thought to mediate interactions with host cells and facilitate parasite invasion, therefore the majority of them are highly expressed in tachyzoites. Micronemal protein 13 (MIC13) is unique in that its expression is low in tachyzoites and is upregulated under bradyzoite-inducing conditions. Previous attempts to disrupt this gene were not successful, implying that it may play critical roles during parasite growth. However, in this study, MIC13 was successfully disrupted in type 1 strain RH and type 2 strain ME49 using CRISPR/Cas9-mediated gene disruption techniques. Consistent with its low expression in tachyzoites and increased expression under stress or bradyzoite-inducing conditions, MIC13-inactivated mutants displayed normal growth, host cell invasion, intracellular replication, and egress, as well as acute virulence at the tachyzoite stage. However, under stress conditions, such as high pH or oxygen limitation, MIC13-disrupted parasites showed significantly slower growth rates compared to the parental strains, suggesting that it is required for optimal parasite growth under bradyzoite-inducing or stress conditions. This is the first micronemal protein reported to have such expression pattern and function modes, which expands our understanding of the diverse functions of micronemal proteins.
研究领域:
寄生虫学
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分子生物学
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传染病学
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JOURNAL OF EXPERIMENTAL BOTANY
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2020
·Vol. 71
·期 19
·5948-5962
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IF 5.908
Deng Changping,Shi Min,Fu Rong,Zhang Yi,Wang Qiang,Zhou Yang,Wang Yao,Ma Xingyuan,Kai Guoyin
Phenolic acids and tanshinones are major bioactive ingredients in Salvia miltiorrhiza, which possess pharmacological activities with great market demand. However, transcriptional regulation of phenolic acid and tanshinone biosynthesis remains poorly understood. Here, a basic leucine zipper transcription factor (TF) named SmbZIP1 was screened from the abscisic acid (ABA)-induced transcriptome library. Overexpression of SmbZIP1 positively promoted phenolic acid biosynthesis by enhancing expression of biosynthetic genes such as cinnamate-4-hydroxylase (C4H1). Furthermore, biochemical experiments revealed that SmbZIP1 bound the G-Box-like1 element in the promoter of the C4H1 gene. Meanwhile, SmbZIP1 inhibited accumulation of tanshinones mainly by suppressing the expression of biosynthetic genes including geranylgeranyl diphosphate synthase (GGPPS) which was confirmed as a target gene by in vitro and in vivo experiments. In contrast, the phenolic acid content was reduced and tanshinone was enhanced in CRISPR/Cas9 [clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9]-mediated knockout lines. In addition, the previously reported positive regulator of tanshinone biosynthesis, SmERF1L1, was found to be inhibited in SmbZIP1 overexpression lines indicated by RNA sequencing, and was proven to be the target of SmbZIP1. In summary, this work uncovers a novel regulator and deepens our understanding of the transcriptional and regulatory mechanisms of phenolic acid and tanshinone biosynthesis, and also sheds new light on metabolic engineering in S. miltiorrhiza.
研究领域:
植物生物学
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分子生物学
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药理学
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基因工程
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PubMed
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Advanced Healthcare Materials
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2025
·
IF 9.6
Liuxian Chen,Qin Huang,Yongjie Liu,Ke Chen,Qing Yang,Hua Tang,Dan Wang,Zhenrong Tang
Complete hepatitis B virus (HBV) cure is hindered primarily by the stable persistence of covalently closed circular DNA (cccDNA). Gene editing approaches to eradicate HBV by targeting cccDNA face challenges and limitations due to suboptimal editing efficiency and substantial off-target effects. Herein, a combinatorial therapeutic strategy is developed that integrates CRISPR/Cas9-mediated cccDNA disruption with an antisense oligonucleotide (ASO)-targeted degradation of pregenomic RNA (pgRNA). To overcome delivery challenges, a hepatocyte-targeting nanocarrier (UACPG) is engineered, featuring low immunogenicity, high payload capacity, and dual-stimuli responsiveness. The UACPG platform enabled liver-specific delivery through surface-conjugated targeting ligands, followed by on-demand release of Cas9 ribonucleoprotein complexes and ASO via RNase H-dependent degradation and near-infrared (NIR) light activation. The results demonstrated that UACPG can effectively reduce HBV replication and viral antigen levels, while significantly lowering cccDNA in hydrodynamic HBV-infected mouse models, with no significant off-target effects observed. This nanocarrier achieved the spatiotemporally controlled release of gene-editing systems in vitro and in vivo, significantly inhibiting the replication of HBV, thereby establishing an innovative technological platform for developing curative HBV therapies.
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Processes
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2026
·Vol. 14
·期 9
·1347
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IF 3.4
Zhuoling Ding,Kadireya Tuerxun,Xueqing Luo,Shishui Zhou
γ-Aminobutyric acid (GABA), a major inhibitory neurotransmitter, is known for its physiological functions in alleviating anxiety and improving sleep. Currently, high-yielding GABA food products are mainly obtained through screening wild-type high-producing strains (e.g.,Saccharomyces cerevisiaeisolated from Sichuan pickles yielding 0.67 g/L) or employing co-culture systems (e.g.,Enterococcus faeciumandLactiplantibacillus plantarumreaching 6.35 g/L). While effective, these methods often rely on natural screening strains or multi-microbial interactions. This study employed CRISPR-Cas9 technology to knockout theUGA1gene inSaccharomyces cerevisiae, a key gene responsible for GABA degradation. Starting from the low higher alcoholSaccharomyces cerevisiaeSY-LH, we successfully constructed the recombinant strain SY-LHU. Remarkably, this study discovered a significant upregulation ofGAD1gene expression followingUGA1knockout, which further enhanced GABA synthesis capacity. Under optimal fermentation conditions (inoculum size 4 × 107cells/mL, wort concentration 10 °P, sugar addition 60 g/L, 30 °C for 10 days, and mixing the malt broth every 48 h), the validation fermentation was performed and the GABA content in the wort beverage reached 280.36 mg/L, representing a 385.4% increase compared to the pre-optimization level. Furthermore, sensory evaluation by a trained panel yielded a mean score of 88, with no significant off-flavors detected, demonstrating the product’s high consumer acceptance. This pioneering work provides a novel and feasible technical pathway for developing functional alcoholic beverages with sleep-aiding properties.
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危险品化学品经营许可证（带存储）