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Cell-state transition can reveal additional information from single-cell ribonucleic acid (RNA)-sequencing data in time-resolved biological phenomena. However, most of the current methods are based on the time derivative of the gene expression state, which restricts them to the short-term evolution of cell states. Here, we present single-cell State Transition Across-samples of RNA-seq data (scSTAR), which overcomes this limitation by constructing a paired-cell projection between biological conditions with an arbitrary time span by maximizing the covariance between two feature spaces using partial least square and minimum squared error methods. In mouse ageing data, the response to stress in CD4+ memory T cell subtypes was found to be associated with ageing. A novel Treg subtype characterized by mTORC activation was identified to be associated with antitumour immune suppression, which was confirmed by immunofluorescence microscopy and survival analysis in 11 cancers from The Cancer Genome Atlas Program. On melanoma data, scSTAR improved immunotherapy-response prediction accuracy from 0.8 to 0.96.
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Perfilação da Expressão Gênica , RNA , Animais , Camundongos , RNA/genética , Perfilação da Expressão Gênica/métodos , Análise de Sequência de RNA/métodos , GenomaRESUMO
Differential expression (DE) gene detection in single-cell ribonucleic acid (RNA)-sequencing (scRNA-seq) data is a key step to understand the biological question investigated. Filtering genes is suggested to improve the performance of DE methods, but the influence of filtering genes has not been demonstrated. Furthermore, the optimal methods for different scRNA-seq datasets are divergent, and different datasets should benefit from data-specific DE gene detection strategies. However, existing tools did not take gene filtering into consideration. There is a lack of metrics for evaluating the optimal method on experimental datasets. Based on two new metrics, we propose single-cell Consensus Optimization of Differentially Expressed gene detection, an R package to automatically optimize DE gene detection for each experimental scRNA-seq dataset.
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RNA , Análise de Célula Única , Perfilação da Expressão Gênica/métodos , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , SoftwareRESUMO
Room-temperature ionic liquids (RTILs) emerged as ideal solvents, and bipyridine as one of the most used ligands have been widely employed in surface science, catalysis, and molecular electronics. Herein, in situ shell-isolated nanoparticle-enhanced Raman spectroscopy (SHINERS) and STM break junction (STM-BJ) technique has been employed to probe the electrochemical process of bipyridine at Au(111)/IL interfaces. It is interestingly found that these molecules undertake a redox process with a pair of well-defined reversible peaks in cyclic voltammograms (CVs). The spectroscopic evidence shows a radical cation generated with rising new Raman peaks related to parallel CC stretching of a positively charged pyridyl ring. Furthermore, these electrochemically charged bipyridine is also confirmed by electrochemical STM-BJ at the single-molecule level, which displays a binary conductance switch ratio of about 400% at the redox potentials. This present work offers a molecular-level insight into the pyridine-mediated reaction process and electron transport in RTILs.
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The CRISPR/Cas9 and Cas12a (Cpf1) tools have been used on a large scale for genome editing. A new effector with a single nuclease domain, a relatively small size, low-frequency off-target effects and cleavage capability under high temperature has been recently established and designated CRISPR/Cas12b (C2c1). Cas12b has also shown temperature inducibility in mammalian systems. Therefore, this system is potentially valuable for editing the genomes of plant species, such as cotton, that are resistant to high temperatures. Using this new system, mutants of upland cotton were successfully generated following Agrobacterium-mediated genetic transformation under a range of temperatures. Transformants (explants infected by Agrobacterium) exposed to 45 °C for 4 days showed the highest editing efficiency. No off-target mutation was detected by whole-genome sequencing. Genome edits by AacCas12b in T0 generation were faithfully passed to the T1 generation. Taken together, CRISPR/Cas12b is therefore an efficient and precise tool for genome editing in cotton plants.
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Sistemas CRISPR-Cas , Edição de Genes , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Genoma de Planta , Gossypium , Temperatura Alta , Humanos , Plantas Geneticamente Modificadas/genética , TetraploidiaRESUMO
Polymerase chain reaction (PCR) and isothermal amplification methods such as LAMP and RPA are widely used for genetic detection. However, there are some shortcomings of these methods such as dependence on thermocycler instruments for PCR, complexity of primer design, the possibility for nonspecific amplification in LAMP and complexity of components in RPA. We develop a novel isothermal DNA detection system named Recombinase Assisted Loop-mediated Amplification (RALA). Recombinase from Thermus thermophilus (TthRecA) was used to open target double-stranded DNA to initiate loop-mediated amplification under isothermal conditions, which simplified the primer design and circumvented pre-denaturation. A FRET sensor named ProofMan and a proofreading enzyme Pfu were introduced to produce fluorescence signals by cleaving the sensor from the 3' end. Consequently, sequence-specific detection based on the RALA system was achieved, and even a single nucleotide polymorphism (SNP) could be identified. By introducing additional loop primers, the fast RALA version can amplify 102 DNA targets in 30 minutes. In addition to high sensitivity and specificity, the flexibility of choosing different reporting sensors makes this method versatile in either quantitative or qualitative DNA detection.
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DNA/análise , Técnicas de Amplificação de Ácido Nucleico/métodos , Recombinases/química , DNA/genética , Primers do DNA/química , Transferência Ressonante de Energia de Fluorescência , Polimorfismo de Nucleotídeo Único , Thermus thermophilus/enzimologiaRESUMO
Riboflavin (vitamin B2) has been thought to be a promising antitumoral agent in photodynamic therapy, though the further application of the method was limited by the unclear molecular mechanism. Our work reveals that riboflavin was able to recognize G-T mismatch specifically and induce single-strand breaks in duplex DNA targets efficiently under irradiation. In the presence of riboflavin, the photo-irradiation could induce the death of tumor cells that are defective in mismatch repair system selectively, highlighting the G-T mismatch as potential drug target for tumor cells. Moreover, riboflavin is a promising leading compound for further drug design due to its inherent specific recognition of the G-T mismatch.
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Pareamento Incorreto de Bases/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Fotoquimioterapia/métodos , Riboflavina/uso terapêutico , Sequência de Bases , Sítios de Ligação/efeitos dos fármacos , Sítios de Ligação/genética , Morte Celular/efeitos dos fármacos , Morte Celular/efeitos da radiação , Dano ao DNA/efeitos dos fármacos , Células HCT116 , Humanos , Luz , Neoplasias/patologia , Riboflavina/farmacologia , Riboflavina/efeitos da radiação , Especificidade por Substrato/efeitos dos fármacosRESUMO
Fluorescence-activated cell sorting (FACS) was exploited to isolate Escherichia coli cells that were highly fluorescent due to the expression of RNA aptamers that induce fluorescence of 3,5-difluoro-4-hydroxybenzylidene imidazolinone. Two different aptamers, named ZT-26 and ZT-324, were identified by this method and compared to the fluorescence-signaling properties of Spinach, a previously reported RNA aptamer. Aptamer ZT-26 exhibits significantly enhanced fluorescence over Spinach only in vitro. However, aptamer ZT-324 is 36% brighter than Spinach when expressed in E. coli. The FACS-based selection strategy presented here is attractive for deriving fluorescent RNA aptamers that function in cells as it directly selects for cells with a high level of fluorescence due to the expression of the RNA aptamer.
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Aptâmeros de Nucleotídeos , Escherichia coli/isolamento & purificação , Citometria de Fluxo/métodos , Proteínas de Fluorescência Verde , Técnica de Seleção de Aptâmeros/métodos , Compostos de Benzil , ImidazolinasRESUMO
Determining correlations between molecules at various levels is an important topic in molecular biology. Large language models have demonstrated a remarkable ability to capture correlations from large amounts of data in the field of natural language processing as well as image generation, and correlations captured from data using large language models can also be applicable to solving a wide range of specific tasks, hence large language models are also referred to as foundation models. The massive amount of data that exists in the field of molecular biology provides an excellent basis for the development of foundation models, and the recent emergence of foundation models in the field of molecular biology has really pushed the entire field forward. We summarize the foundation models developed based on RNA sequence data, DNA sequence data, protein sequence data, single-cell transcriptome data, and spatial transcriptome data respectively, and further discuss the research directions for the development of foundation models in molecular biology.
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BACKGROUND: Diabetes is a global health problem whose common complication is diabetic cardiomyopathy, characterized by chronic inflammation of the heart muscle. Macrophages are the main white blood cells found in the resting heart. Therefore, we investigated the underling mechanism of macrophage on myocardial fibrosis in diabetes. METHODS: Here, echocardiography was utilized to evaluate cardiac function, and the degree of myocardial fibrosis was assessed using Masson's trichrome staining, followed by single-cell RNA sequencing (scRNA-seq) to analyze the phenotype, function, developmental trajectory, and interactions between immune cells, endothelial cells (ECs), and fibroblasts (FBs) in the hearts of db/db mice at different stages of diabetes. Macrophages and cardiac fibroblasts were also co-cultured in order to study the signaling between macrophages and fibroblasts. RESULTS: We found that with the development of diabetes mellitus, myocardial hypertrophy and fibrosis occurred that was accompanied by cardiac dysfunction. A significant proportion of immune cells, endothelial cells, and fibroblasts were identified by RNA sequencing. The most significant changes observed were in macrophages, which undergo M1 polarization and are critical for oxidative stress and extracellular matrix (ECM) formation. We further found that M1 macrophages secreted interleukin-1ß (IL-1ß), which interacted with the receptor on the surface of fibroblasts, to cause myocardial fibrosis. In addition, crosstalk between M1 macrophages and endothelial cells also plays a key role in fibrosis and immune response regulation through IL-1ß and corresponding receptors. CONCLUSIONS: M1 macrophages mediate diabetic myocardial fibrosis through interleukin-1ß interaction with fibroblasts.
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Diabetes Mellitus , Cardiomiopatias Diabéticas , Camundongos , Animais , Interleucina-1beta , Células Endoteliais , Macrófagos , FibroseRESUMO
Molybdenum and its alloys are known for their superior strength among body-centered cubic materials. However, their widespread application is hindered by a significant decrease in ductility at lower temperatures. In this study, we demonstrate the achievement of exceptional ductility in a Mo alloy containing rare-earth La2O3 nanoparticles through rotary-swaging, a rarity in Mo-based materials. Our analysis reveals that the large ductility originates from substantial variations in the electronic density of states, a characteristic intrinsic to rare-earth elements. This characteristic can accelerate the generation of oxygen vacancies, facilitating the amorphization of the oxide-matrix interface. This process promotes vacancy absorption and modification of dislocation configurations. Furthermore, by inducing irregular shapes in the La2O3 nanoparticles through rotary-swaging, incoming dislocations interact with them, creating multiple dislocation sources near the interface. These dislocation sources act as potent initiators at even reduced temperatures, fostering diverse dislocation types and intricate networks, ultimately enhancing dislocation plasticity.
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Polymer materials with multiple stimuli-responsive properties have demonstrated many potential and practical applications. By covalently introducing spiropyran (SP1) and spirothiopyran (STP) into the polyurethane backbone, photochromic, mechanochromic, and thermally discolored polymer materials have been prepared. In this work, we report for the first time that white light (violet, blue, and green light) above a certain intensity can activate STP to green color. Based on the above discovery, the polyurethane with SP1 and STP can exhibit reversible three-color changes (brown, green, and purple) in response to four stimuli: ultraviolet irradiation, white light irradiation, mechanical stress, and heat. The color-changing polymer materials have high color contrast and excellent reversibility, and can be used for reversible writing, anticounterfeiting and information encryption, etc.
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Magnesium and its alloys are considered excellent materials for biodegradable implants because of their good biocompatibility and biodegradability as well as their mechanical properties. However, the rapid degradation rate severely limits their clinical applications. Plasma electrolytic oxidation (PEO), also known as micro-arc oxidation (MAO), is an effective surface modification technique. However, there are many pores and cracks on the coating surface under conventional PEO process. The corrosive products tend to penetrate deeply into the substrate, reducing its corrosion resistance and the biocompatibility, which makes PEO-coated Mg difficult to meet the long-term needs of in vivo implants. Hence, it is necessary to modify the PEO coating. This review discusses the formation mechanism and the influential parameters of PEO coatings on Mg. This is followed by a review of the latest research of the pretreatment and typical amelioration of PEO coating on biodegradable Mg alloys in the past 5 years, including calcium phosphate (Ca-P) coating, layered double hydroxide (LDH)-PEO coating, ZrO2 incorporated-PEO coating, antibacterial ingredients-PEO coating, drug-PEO coating, polymer-PEO composite coating, Plasma electrolytic fluorination (PEF) coating and self-healing coating. Meanwhile, the improvements of morphology, corrosion resistance, wear resistance, biocompatibility, antibacterial abilities, and drug loading abilities and the preparation methods of the modified PEO coatings are deeply discussed as well. Finally, the challenges and prospects of PEO coatings are discussed in detail for the purpose of promoting the clinical application of biodegradable Mg alloys.
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An environmentally friendly corrosion inhibitor was prepared from the bio-based platform 5-hydroxymethylfurfural. This corrosion inhibitor was confirmed to be an efficient mixed-type corrosion inhibitor through a weight loss experiment and electrochemical experiment. Both thermodynamic and kinetic parameters were calculated and discussed, indicating that the adsorption of this bio-based inhibitor on a steel surface is a chemisorption process. Moreover, quantum chemical calculations were performed and further confirmed the formation of an effective productive film of this bio-based inhibitor on the metal surface. It is worth noting that the synthesis route of this bio-based corrosion inhibitor is green and environmentally friendly, and does not involve toxic chemical reagents.
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Internal strain and its distribution within the crystal lattice play crucial roles in modulating dislocation activities, thereby affecting mechanical properties of materials. Through the synergistic application of integrated differential phase contrast, in situ transmission electron microscopy characterizations, and computational simulations, a method is unveiled for homogenizing dislocation pinning in NiCoCr multi-principal element alloy (MPEA) through the introduction of a high concentration of oxygen atoms with high diffusion mobility. The doping of massive oxygen atoms creates a high density of strong local pinning points for dislocation motion. Notably, oxygen interstitials exhibit remarkable diffusion and mobility across different octahedral and tetrahedral sites within the distorted crystal lattice of NiCoCrO alloy, even at room temperature. The capability allows for the release of severe stress concentrations arising from dislocation entanglement and the establishment of new strong local pinning points at alternative locations in a uniform way, enabling the material with high strength and outstanding deformability. These findings suggest that interstitial atoms can exhibit significant mobility, even at ambient temperature, in complex MPEAs with spreading lattice distortion, opening new possibilities for dislocation engineering.
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Insects pose significant challenges in cotton-producing regions. Here, they describe a high-throughput CRISPR/Cas9-mediated large-scale mutagenesis library targeting endogenous insect-resistance-related genes in cotton. This library targeted 502 previously identified genes using 968 sgRNAs, generated ≈2000 T0 plants and achieved 97.29% genome editing with efficient heredity, reaching upto 84.78%. Several potential resistance-related mutants (10% of 200 lines) their identified that may contribute to cotton-insect molecular interaction. Among these, they selected 139 and 144 lines showing decreased resistance to pest infestation and targeting major latex-like protein 423 (GhMLP423) for in-depth study. Overexpression of GhMLP423 enhanced insect resistance by activating the plant systemic acquired resistance (SAR) of salicylic acid (SA) and pathogenesis-related (PR) genes. This activation is induced by an elevation of cytosolic calcium [Ca2+ ]cyt flux eliciting reactive oxygen species (ROS), which their demoted in GhMLP423 knockout (CR) plants. Protein-protein interaction assays revealed that GhMLP423 interacted with a human epidermal growth factor receptor substrate15 (EPS15) protein at the cell membrane. Together, they regulated the systemically propagating waves of Ca2+ and ROS, which in turn induced SAR. Collectively, this large-scale mutagenesis library provides an efficient strategy for functional genomics research of polyploid plant species and serves as a solid platform for genetic engineering of insect resistance.
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Sistemas CRISPR-Cas , RNA Guia de Sistemas CRISPR-Cas , Humanos , Animais , Sistemas CRISPR-Cas/genética , Espécies Reativas de Oxigênio/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , InsetosRESUMO
Osteoarthritis (OA) is one of the most common diseases in the orthopedic clinic, characterized by progressive cartilage degradation. RNA-binding proteins (RBPs) are capable of binding to RNAs at transcription and translation levels, playing an important role in the pathogenesis of OA. This study aims to investigate the diagnosis values of RBP-related genes in OA. The RBPs were collected from previous studies, and the GSE114007 dataset (control = 18, OA = 20) was downloaded from the Gene Expression Omnibus (GEO) as the training cohort. Through various bioinformatical and machine learning methods, including genomic difference detection, protein-protein interaction network analyses, Lasso regression, univariate logistic regression, Boruta algorithm, and SVM-RFE, RNMT and RBM24 were identified and then included into the random forest (RF) diagnosis model. GSE117999 dataset (control = 10, OA = 10) and clinical samples collected from local hospital (control = 10, OA = 11) were used for external validation. The RF model was a promising tool to diagnose OA in the training dataset (area under curve [AUC] = 1.000, 95% confidence interval [CI] = 1.000-1.000), the GSE117999 cohort (AUC = 0.900, 95% CI = 0.769-1.000), and local samples (AUC = 0.759, 95% CI = 0.568-0.951). Besides, qPCR and Western Blotting experiments showed that RNMT (P < 0.05) and RBM24 (P < 0.01) were both down-regulated in CHON-001 cells with IL-1ß treatment. In all, an RF model to diagnose OA based on RNMT and RBM24 in cartilage tissue was constructed, providing a promising clinical tool and possible cut-in points in molecular mechanism clarification.
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Condrócitos , Osteoartrite , Humanos , Condrócitos/metabolismo , Algoritmo Florestas Aleatórias , Cartilagem/metabolismo , Osteoartrite/metabolismo , Articulação do Joelho , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
Alkyl chain modification strategies in both organic semiconductors and inorganic dielectrics play a crucial role in improving the performance of organic thin-film transistors (OTFTs). Polyimide (PI) and its derivatives have received extensive attention as dielectrics for application in OTFTs because of flexibility, high-temperature resistance, and low cost. However, low-temperature solution processing PI-based gate dielectric for flexible OTFTs with high mobility, low operating voltage, and high operational stability remains an enormous challenge. Furthermore, even though di-n-decyldinaphtho[2,3-b:2',3'-f]thieno[3,2-b]thiophene (C10-DNTT) is known to have very high mobility as an air-stable and high-performance organic semiconductor, the C10-DNTT-based TFTs on the PI gate dielectrics still showed relatively low mobility. Here, inspired by alkyl side chain engineering, we design and synthesize a series of PI materials with different alkyl side chain lengths and systematically investigate the PI surface properties and the evolution of organic semiconductor morphology deposited on PI surfaces during the variation of alkyl side chain lengths. It is found that the alkyl side chain length has a critical influence on the PI surface properties, as well as the grain size and molecular orientation of semiconductors. Good field-effect characteristics are obtained with high mobilities (up to 1.05 and 5.22 cm2/Vs, which are some of the best values reported to date), relatively low operating voltage, hysteresis-free behavior, and high operational stability in OTFTs. These results suggest that the strategy of optimizing alkyl side-chain lengths opens up a new research avenue for tuning semiconductor growth to enable high mobility and outstanding operational stability of PI-based OTFTs.
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Introduction: The clubroot disease caused by Plasmodiophora brassicae (P. brassicae) poses a serious threat to the economic value of cruciferous crops, which is a serious problem to be solved worldwide. Some resistance genes to clubroot disease in Brassica rapa L. ssp pekinensis cause by P. brassicae have been located on different chromosomes. Among them, Rcr1 and Rcr2 were mapped to the common candidate gene Bra019410, but its resistance mechanism is not clear yet. Methods: In this experiment, the differences of BrRPP1 between the resistant and susceptible material of Chinese cabbage were analyzed by gene cloning and qRT-PCR. The gene function was verified by Arabidopsis homologous mutants. The expression site of BrRPP1 gene in cells was analyzed by subcellular localization. Finally, the candidate interaction protein of BrRPP1 was screened by yeast two-hybrid library. Results: The results showed that the cDNA sequence, upstream promoter sequence and expression level of BrRPP1 were quite different between the resistant and susceptible material. The resistance investigation found that the Arabidopsis mutant rpp1 was more susceptible to clubroot disease than the wild type, which suggested that the deletion of rpp1 reduces resistance of plant to clubroot disease. Subcellular location analysis confirmed that BrRPP1 was located in the nucleus. The interaction proteins of BrRPP1 screened from cDNA Yeast Library by yeast two-hybrid are mainly related to photosynthesis, cell wall modification, jasmonic acid signal transduction and programmed cell death. Discussion: BrRPP1 gene contains TIR-NBS-LRR domain and belongs to R gene. The cDNA and promoter sequence of BrRPP1 in resistant varieties was different from that in susceptible varieties led to the significant difference of the gene expression of BrRPP1 between the resistant varieties and the susceptible varieties. The high expression of BrRPP1 gene in resistant varieties enhanced the resistance of Chinese cabbage to P. brassicae, and the interaction proteins of BrRPP1 are mainly related to photosynthesis, cell wall modification, jasmonic acid signal transduction and programmed cell death. These results provide important clues for understanding the mechanism of BrRPP1 in the resistance of B. rapa to P. brassicae.
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A deep understanding of immunotherapy response/resistance mechanisms and a highly reliable therapy response prediction are vital for cancer treatment. Here, we developed scCURE (single-cell RNA sequencing [scRNA-seq] data-based Changed and Unchanged cell Recognition during immunotherapy). Based on Gaussian mixture modeling, Kullback-Leibler (KL) divergence, and mutual nearest-neighbors criteria, scCURE can faithfully discriminate between cells affected or unaffected by immunotherapy intervention. By conducting scCURE analyses in melanoma and breast cancer immunotherapy scRNA-seq data, we found that the baseline profiles of specific CD8+ T and macrophage cells (identified by scCURE) can determine the way in which tumor microenvironment immune cells respond to immunotherapy, e.g., antitumor immunity activation or de-activation; therefore, these cells could be predictive factors for treatment response. In this work, we demonstrated that the immunotherapy-associated cell-cell heterogeneities revealed by scCURE can be utilized to integrate the therapy response mechanism study and prediction model construction.