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Proximity labeling has emerged as a powerful strategy for interrogating cell-cell interactions. However, the nanometer-scale labeling radius impedes the use of current methods for indirect cell communications and makes recording cell spatial organization in tissue samples difficult. Here, we develop quinone methide–assisted identification of cell spatial organization (QMID), a chemical strategy with the labeling radius matching the cell dimension. The activating enzyme is installed on the surface of bait cells, which produces QM electrophiles that can diffuse across micrometers and label proximal prey cells independent of cell-cell contacts. In cell coculture, QMID reveals gene expression of macrophages that are regulated by spatial proximity to tumor cells. Furthermore, QMID enables labeling and isolation of proximal cells of CD4 + and CD8 + T cells in the mouse spleen, and subsequent single-cell RNA sequencing uncovers distinctive cell populations and gene expression patterns within the immune niches of specific T cell subtypes. QMID should facilitate dissecting cell spatial organization in various tissues.

Clickable unnatural sugars have been widely used in studying glycosylation in living systems via the metabolic glycan labelling (MGL) strategy. Partial protection of unnatural sugars by 1,6-di-O-acylation increases the labelling efficiency while avoiding the non-specific S-glyco-modification. Herein, we report the facile synthesis of a series of clickable unnatural sugars in both the unprotected and 1,6-di-O-acylated forms at the ten-gram scale. By evaluation of the labelling specificity, efficiency, and biocompatibility of various 1,6-di-O-acylated sugars for MGL in cell lines and living mice, we demonstrate that 1,6-di-O-propionylated unnatural sugars are optimal chemical reporters for glycan labelling. The synthetic routes developed in this work should facilitate the widespread use of MGL with no artificial S-glyco-modification for investigating the functional roles of glycans.

Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful tool for profiling gene expression of distinct cell populations at the single-cell level. However, the information of the positions of cells within the multicellular samples is missing in scRNA-seq datasets. To overcome this limitation, we herein develop OpTAG (optical cell tagging) as a new chemical platform for attaching functional tags onto cell surfaces in a spatially resolved manner. With OpTAG, we establish OpTAG-seq, which enables spatially resolved scRNA-seq. We apply OpTAG-seq to investigate the spatially defined transcriptional program in migrating cancer cells and identified a list of genes that are potential regulators for cancer cell migration and invasion. OpTAG-seq provides a convenient method for mapping cellular heterogeneity with spatial information within multicellular biological systems.

Interferon-regulatory factor 5 (IRF5) participates in the regulation of apoptosis, affects the phenotype of inflammatory macrophages and plays an essential role in the inflammatory response. However, the role of IRF5 in the progression of amyotrophic lateral sclerosis (ALS) remains largely unknown. Here, we show that IRF5 mainly accumulated in the nucleus in cells expressing the truncated 25 k

Simple Summary The spread of citrus Huanglongbing caused significant damage to the world’s citrus industry. Thermotherapy and chemical agents were used to control this disease; however, the effectiveness of these treatments is frequently inconsistent. In addition, CLas cannot be cultured in vitro. Therefore, structure-based virtual screening is a novel method to find compounds that work against CLas. This study used CLas GMPS as a target for high-throughput screening and selected some compounds which have a higher binding affinity to test their inhibition of CLas GMPS. Finally, two molecules were identified as the lead compound to control citrus HLB. Abstract Citrus production is facing an unprecedented problem because of huanglongbing (HLB) disease. Presently, no effective HLB-easing method is available when citrus becomes infected. Guanosine 5′-monophosphate synthetase (GMPS) is a key protein in the de novo synthesis of guanine nucleotides. GMPS is used as an attractive target for developing agents that are effective against the patogen infection. In this research, homology modeling, structure-based virtual screening, and molecular docking were used to discover the new inhibitors against CLas GMPS. Enzyme assay showed that folic acid and AZD1152 showed high inhibition at micromole concentrations, with AZD1152 being the most potent molecule. The inhibition constant (Ki) value of folic acid and AZD1152 was 51.98 µM and 4.05 µM, respectively. These results suggested that folic acid and AZD1152 could be considered as promising candidates for the development of CLas agents.

Citrus huanglongbing (HLB) is a destructive disease caused by Candidatus Liberibacter species and is a serious global concern for the citrus industry. To date, there is no established strategy for control of this disease. Previously, Bacillus amyloliquefaciens GJ1 was screened as the biocontrol agent against HLB. In this study, two-year-old citrus infected by Ca. L. asiaticus were treated with B. amyloliquefaciens GJ1 solution via root irrigation. In these plants, after seven irrigation treatments, the results indicated that the photosynthetic parameters, chlorophyll content, resistance-associated enzyme content and the expression of defense-related genes were significantly higher than for the plants treated with the same volume water. The content of starch and soluble sugar were significantly lower, compared to the control treatment. The parallel reaction monitoring (PRM) results revealed that treatment with B. amyloliquefaciens GJ1 solution, the expression levels of 3 proteins with photosynthetic function were upregulated in citrus leaves. The accumulation of reactive oxygen species (ROS) in citrus leaves treated with B. amyloliquefaciens GJ1 flag22 was significantly higher than untreated plants and induced the defense-related gene expression in citrus. Finally, surfactin was identified from the fermentation broth of B. amyloliquefaciens GJ1 by high-performance liquid chromatography. These results indicate that B. amyloliquefaciens GJ1 may improve the immunity of citrus by increasing the photosynthesis and enhancing the expression of the resistance-related genes.

Background: An ideal animal model to study SARS-coronavirus 2 (SARS-CoV-2) pathogenesis and evaluate therapies and vaccines should reproduce SARS-CoV-2 infection and recapitulate lung disease like those seen in humans. The angiotensin-converting enzyme 2 (ACE2) is a functional receptor for SARS-CoV-2, but mice are resistant to the infection because their ACE2 is incompatible with the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. Methods: We generated a mouse-adapted strain SARS-CoV-2 by serial passages in the lung of BALB/c mice. Complete genome deep sequencing of different generations of viruses was performed to characterize the dynamics of the adaptive mutations in SARS-CoV-2. Indirect immunofluorescence analysis and Biolayer interferometry experiments demonstrated that two mutations in RBD significantly increased its binding affinity towards mouse ACE2. Significantly, TLR7/8 agonist Resiquimod block SARS-CoV-2 in vitro and in vivo. Findings: We adapted a wild-type SARS-CoV-2 by serial passages in the lung of BALB/c mice. The mouse-adapted strain WBP-1 showed increased infectivity in BALB/c mice and led to severe interstitial pneumonia. We characterized the dynamics of the adaptive mutations in SARS-CoV-2 and demonstrated that Q493K and Q498H in RBD significantly increased its binding affinity towards mouse ACE2. Additionally, The TLR7/8 agonist Resiquimod was able to protect mice against WBP-1 challenge, demonstrating this mouse-adapted strain is a useful tool to investigate COVID-19 and develop new therapies. Interpretation: We found for the first time that the Q493K and Q498H mutations in the RBD of WBP-1 enhanced its interactive affinities with mACE2. The mouse-adapted SARS-CoV-2 provides a valuable tool for the evaluation of novel antiviral and vaccine strategies, especially in determining the immunopathological consequences of any intervention. This study also verified the antiviral activity of TLR7/8 agonist Resiquimod against SARS-CoV-2 in vitro and in vivo. Funding Statement: This research was funded by Emergency Science and Technology Project of Hubei Province（2020FCA046）and Independent Science and Technology Innovation Fund of Huazhong Agricultural University in 2020 (2662020PY002). Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: The animal experiments were approved by the Research Ethics Committee, Huazhong Agricultural University, Hubei, China (HZAUMO-2020-0007). All the animal experiments were conducted in accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals from the Research Ethics Committee, Huazhong Agricultural University, Hubei, China.

Citrus huanglongbing (HLB) is a destructive disease caused by Candidatus Liberibacter species and is a serious global concern for the citrus industry. To date, there is no established strategy for control of this disease. Previously, Bacillus amyloliquefaciens GJ1 was screened as the biocontrol agent against HLB. In this study, two-year-old citrus infected by Ca. L. asiaticus were treated with B. amyloliquefaciens GJ1 solution via root irrigation. In these plants, after seven irrigation treatments, the results indicated that the photosynthetic parameters, chlorophyll content, resistance-associated enzyme content and the expression of defense-related genes were significantly higher than for the plants treated with the same volume water. The content of starch and soluble sugar were significantly lower, compared to the control treatment. The parallel reaction monitoring (PRM) results revealed that treatment with B. amyloliquefaciens GJ1 solution, the expression levels of 3 proteins with photosynthetic function were upregulated in citrus leaves. The accumulation of reactive oxygen species (ROS) in citrus leaves treated with B. amyloliquefaciens GJ1 flag22 was significantly higher than untreated plants and induced the defense-related gene expression in citrus. Finally, surfactin was identified from the fermentation broth of B. amyloliquefaciens GJ1 by high-performance liquid chromatography. These results indicate that B. amyloliquefaciens GJ1 may improve the immunity of citrus by increasing the photosynthesis and enhancing the expression of the resistance-related genes.

Cyanobacterial blooms have a range of social, environmental and economic impacts due to their content of secondary metabolites involving toxins. Microcystins (MCs) is one of the cyanotoxins found in freshwater ecosystems. This study was aimed to adopt a rapid technique for determining the microcystin in blooms and freshwater by the detection of phycocyanin operon of cyanobacteria using PCβF and PCαR primers as well as mcyE gene encoding to microcystin/ nodularin using HEP primers. The molecular results showed that phycocyanin operon and mcyE gene were disclosed in the freshwater and bloom of samples at the studied sites. In conclusion, The PCR assay used in this study was helpful and rapid, in particular when the target organism concentration in the freshwater sample is very low.

Background An ideal animal model to study SARS-coronavirus 2 (SARS-CoV-2) pathogenesis and evaluate therapies and vaccines should reproduce SARS-CoV-2 infection and recapitulate lung disease like those seen in humans. The angiotensin-converting enzyme 2 (ACE2) is a functional receptor for SARS-CoV-2, but mice are resistant to the infection because their ACE2 is incompatible with the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein . Methods SARS-CoV-2 was passaged in BALB/c mice to obtain mouse-adapted virus strain. Complete genome deep sequencing of different generations of viruses was performed to characterize the dynamics of the adaptive mutations in SARS-CoV-2. Indirect immunofluorescence analysis and Biolayer interferometry experiments determined the binding affinity of mouse-adapted SARS-CoV-2 WBP-1 RBD to mouse ACE2 and human ACE2. Finally, we tested whether TLR7/8 agonist Resiquimod (R848) could also inhibit the replication of WBP-1 in the mouse model. Findings The mouse-adapted strain WBP-1 showed increased infectivity in BALB/c mice and led to severe interstitial pneumonia. We characterized the dynamics of the adaptive mutations in SARS-CoV-2 and demonstrated that Q493K and Q498H in RBD significantly increased its binding affinity towards mouse ACE2. Additionally, the study tentatively found that the TLR7/8 agonist Resiquimod was able to protect mice against WBP-1 challenge. Therefore, this mouse-adapted strain is a useful tool to investigate COVID-19 and develop new therapies. Interpretation We found for the first time that the Q493K and Q498H mutations in the RBD of WBP-1 enhanced its interactive affinities with mACE2. The mouse-adapted SARS-CoV-2 provides a valuable tool for the evaluation of novel antiviral and vaccine strategies. This study also tentatively verified the antiviral activity of TLR7/8 agonist Resiquimod against SARS-CoV-2 in vitro and in vivo. Funding This research was funded by the National Key Research and Development Program of China (2020YFC0845600) and Emergency Science and Technology Project of Hubei Province (2020FCA046) and Robert A. Welch Foundation (C-1565).

Citrus huanglongbing (HLB) is a destructive disease that causes significant damage to many citrus producing areas worldwide. To date, no strategy against this disease has been established. Inosine 5&prime, monophosphate dehydrogenase (IMPDH) plays crucial roles in the de novo synthesis of guanine nucleotides. This enzyme is used as a potential target to treat bacterial infection. In this study, the crystal structure of a deletion mutant of CLas IMPDH&Delta, 98-201 in the apo form was determined. Eight known bioactive compounds were used as ligands for molecular docking. The results showed that bronopol and disulfiram bound to CLas IMPDH&Delta, 98-201 with high affinity. These compounds were tested for their inhibition against CLas IMPDH&Delta, 98-201 activity. Bronopol and disulfiram showed high inhibition at nanomolar concentrations, and bronopol was found to be the most potent molecule (Ki = 234 nM). The Ki value of disulfiram was 616 nM. These results suggest that bronopol and disulfiram can be considered potential candidate agents for the development of CLas inhibitors.

This paper studies an audio similarity comparison system on Android platform to judge the similarity of dubbing audio in some contexts. The system mainly uses the image similarity comparison method and the MFCC algorithm as its algorithm basis. Considering the limitation of mobile platform, the current mobile audio comparison methods are studied and improved, to reduce operation time and improve accuracy of the comparison.

Rationale: Cardiac fatty acid-binding protein (FABP3) is a cardiac-specific member of lipid-binding protein family and its expression level is often reduced in patients with diabetic heart diseases. Recent evidence suggests that FABP3 suppresses calcium transient and shortening of isolated rat cardiomyocytes. However, the underlying mechanisms are largely elusive. Objective: To determine the role of FABP3 in regulating SR calcium release. Results: In STZ-induced type I diabetes mouse model (DM), protein expression level of FABP3 was elevated, cardiac function was reduced and negatively correlated with FABP3 expression level. Amplitudes of cell shortening and calcium transient are both impaired in DM cardiomyocytes, which can be mimicked by applying FABP3 at pathological concentration to control cardiomyocytes. FABP3 reduces calcium transient amplitude of cardiomyocytes in a dose dependent manner (EC50 = 0.052 nmol/L). SR calcium content is reduced in DM cardiomyocytes and FABP3 depletes SR calcium content in cardiomyocytes. FABP3 colocalizes with SERCA, inhibits SERCA activity with a greater EC50 (0.49 nmol/L). Co-immunoprecipitation study suggests elevated FABP3 promotes binding between SERCA and phospholamban. FABP3 also colocalizes with RyR2, binds to RyR2 and promotes RyR2-mediated SR calcium leak. Conclusion: Increased FABP3 expression level in DM mice compromises cardiac function by reducing SR calcium load via two independent approaches: (A) to reduce SERCA activity by promoting PLB-SERCA interaction and (B) to enhance RyR2-mediated SR calcium leak.