Advances in miniature CRISPR-Cas proteins and their applications in gene editing.

The CRISPR-Cas system consists of Cas proteins and single-stranded RNAs that recruit Cas proteins and specifically target the nucleic acid. Some Cas proteins can accurately cleave the target nucleic acid under the guidance of the single-stranded RNAs. Due to its exceptionally high specificity, the CRISPR-Cas system is now widely used in various fields such as gene editing, transcription regulation, and molecular diagnosis. However, the huge size of the most frequently utilized Cas proteins (Cas9, Cas12a, and Cas13, which contain 950-1,400 amino acids) can limit their applicability, especially in eukaryotic gene editing, where larger Cas proteins are difficult to deliver into the target cells. Recently discovered miniature CRISPR-Cas proteins, consisting of only 400 to 800 amino acids, offer the possibility of overcoming this limitation. This article systematically reviews the latest research progress of several miniature CRISPR-Cas proteins (Cas12f, Cas12j, Cas12k, and Cas12m) and their practical applications in the field of gene editing.

Design, synthesis and bioactivity evaluation of triazole antifungal drugs with phenylthiophene structure.

Invasive fungal infections have high morbidity and mortality rates and have become one of the most serious threats to human health. In the present study, a series of triazole antifungal derivatives with phenylthiophene backbone were obtained by structural modification of the lead compound using Iodiconazole as the lead compound. Among them, compound 19g is a triazole antifungal compound with 4-chloro-2-fluoro phenylthiophene backbone, which showed optimal antifungal activity against Candida albicans, Cryptococcus neoformans, and Aspergillus, with a MIC80 value of 0.0625 µg/mL. In addition, compounds 19e, 19f, 19g, 19h, 19i and 19k exhibited different levels of inhibitory activity against fluconazole-resistant strains with MIC80 values ranging from 0.0625 µg/mL to 32 µg/mL. Since compound 19g had optimal in vitro antifungal activity, we selected 19g for human liver microsomal stability and CYP enzyme inhibition assays as well as further evaluated the inhibitory activity of compound 19g on normal and cancerous cells in humans. Finally, we verified the inhibitory effect of compound 19g on the filamentation of Candida albicans and determined the mechanism of action by sterol composition analysis.

Pursuing sustainable productivity with millions of smallholder farmers.

Sustainably feeding a growing population is a grand challenge, and one that is particularly difficult in regions that are dominated by smallholder farming. Despite local successes, mobilizing vast smallholder communities with science- and evidence-based management practices to simultaneously address production and pollution problems has been infeasible. Here we report the outcome of concerted efforts in engaging millions of Chinese smallholder farmers to adopt enhanced management practices for greater yield and environmental performance. First, we conducted field trials across China's major agroecological zones to develop locally applicable recommendations using a comprehensive decision-support program. Engaging farmers to adopt those recommendations involved the collaboration of a core network of 1,152 researchers with numerous extension agents and agribusiness personnel. From 2005 to 2015, about 20.9 million farmers in 452 counties adopted enhanced management practices in fields with a total of 37.7 million cumulative hectares over the years. Average yields (maize, rice and wheat) increased by 10.8-11.5%, generating a net grain output of 33 million tonnes (Mt). At the same time, application of nitrogen decreased by 14.7-18.1%, saving 1.2 Mt of nitrogen fertilizers. The increased grain output and decreased nitrogen fertilizer use were equivalent to US$12.2 billion. Estimated reactive nitrogen losses averaged 4.5-4.7 kg nitrogen per Megagram (Mg) with the intervention compared to 6.0-6.4 kg nitrogen per Mg without. Greenhouse gas emissions were 328 kg, 812 kg and 434 kg CO2 equivalent per Mg of maize, rice and wheat produced, respectively, compared to 422 kg, 941 kg and 549 kg CO2 equivalent per Mg without the intervention. On the basis of a large-scale survey (8.6 million farmer participants) and scenario analyses, we further demonstrate the potential impacts of implementing the enhanced management practices on China's food security and sustainability outlook.

Knockdown of CPSF4 Inhibits Bladder Cancer Cell Growth by Upregulating NRF1.

Increasing studies have shown that nuclear respiratory factor 1 (NRF1) deficiency frequently occurs in many human diseases, and its activation can protect neurons and other cells from degenerative diseases and malignant tumors. However, how NRF1 is regulated in bladder cancer remains unknown. Our research aims to reveal the role of leavage and polyadenylation-specific factor 4 (CPSF4) on the growth inhibition effect of bladder cancer and clarify its relationship with NRF1. Here, cell proliferation assay, transwell migration assay and multicellular tumor spheroids (MCTS) formation assay in the bladder cancer cell lines were carried out to measure tumor cell growth. Western bolt assay was carried out to identify the relationship between NRF1 and CPSF4. Also, subcutaneous xenograft tumors in nude mice were established to further validate the inhibition effect of CPSF4 on bladder tumor and the regulation on NRF1. The results in vitro showed that knockdown of CPSF4 strongly reduced the proliferation and migration, and inhibited MCTS formation in 5637 and HT1376 cell lines, while an additional knockdown of increased NRF1 induced by CPSF4 knockdown partially abolished these effects. The results in vivo showed that knockdown of CPSF4 strongly reduced the volume and weight of subcutaneous tumor, and decreased the expression of Ki-67 in tumor tissue, while NRF1 knockdown partially reversed these effects induced by CPSF4 knockdown. Western bolt assay demonstrated that CPSF4 could negatively regulate NRF1. Our results indicated that knock-down of CPSF4 inhibited bladder cancer cell growth by upregulating NRF1, which might provide evidence of CPSF4 as a therapeutic target for bladder cancer.

Design, synthesis, and biological evaluation of a potent PLK4 inhibitor WY29 with 1H-pyrazolo[3,4-d]pyrimidine scaffold.

Centriole duplication occurs once per cell cycle and is regulated by Polo-like kinase 4 (PLK4). Overexpression of PLK4 in somatic cells can lead to the excessive formation of centrioles, directly causing chromosome segregation errors and tumorigenesis. In this study, we described our efforts to develop a series of PLK4 inhibitors with 1H-pyrazolo[3,4-d]pyrimidine core, and further structure- and receptor-based design and optimization resulted in a potent inhibitor WY29 (IC50 = 0.027 µM), which exhibited good selectivity to other PLK family members (PLK1-3). At the cellular level, compound WY29 showed excellent antiproliferative activity against three breast cancer cell lines (MCF-7, BT474, and MDA-MB-231) while weak inhibitory activity was found on normal cell line HUVECs. In addition, the in vitro preliminary drug-like properties evaluation of compound WY29 showed outstanding stability in human plasma and liver microsomes, and weak inhibitory activity against the major subtypes of human cytochrome P450. Also, the drug-like properties prediction of compound WY29 displayed remarkable drug-like properties (drug-likeness mode score: 1.06). In conclusion, these results support the further development of compound WY29 as a lead compound for PLK4-targeted anticancer drug discovery.

Design, synthesis, and biological evaluation of aminopyridine derivatives as novel tropomyosin receptor kinase inhibitors.

Tropomyosin receptor kinase (TRK) is a successful target for the treatment of various cancers caused by NTRK gene fusions. Herein, based on a rational drug design strategy, we designed and synthesized 35 aminopyrimidine derivatives that were shown to be TRKA inhibitors in the enzyme assay, among which compounds C3, C4, and C6 showed potent inhibitory activities against TRKA with IC50 values of 6.5, 5.0, and 7.0 nM, respectively. In vitro antiproliferative activity study showed that compound C3 significantly inhibited the proliferation of KM-12 cells but had weak inhibitory effect on MCF-7 cells and HUVEC cells. The preliminary druggability evaluation showed that compound C3 exhibited favorable liver microsomal and plasma stabilities and had weak or no inhibitory activity against cytochrome P450 isoforms at 10 µM. Compounds C3, C4, and C6 were also selected for ADME (absorption, distribution, metabolism, and elimination) properties prediction and molecular docking studies. Inhibition experiments showed that compound C3 was not selective for TRK subtypes. All results indicated that compound C3 was a useful candidate for the development of TRK inhibitors.

Amendment with controlled release urea increases leaf morpho-physiological traits, grain yield and NUE in a double-cropping rice system in southern China.

BACKGROUND: Understanding of mechanisms that underpin high-yielding cropping systems is essential for optimizing management practices. Currently, the contribution of plant traits such as leaf area, chlorophyll content and intercepted photosynthetically active radiation (PARi ) to yield and nitrogen use efficiency (NUE) are not fully understood. In addition, the understanding of how canopy traits are affected by nitrogen (N) management practices is unclear. The present study aimed to determine the effect of amendment with controlled release urea (CR), common urea or no urea on NUE and plant eco-physiological characteristics in a 2-year field study in a double rice cropping system. RESULTS: Regulation of N release through amendment with CR significantly increased grain yield, NUE and leaf morpho-physiological attributes. CR coupled with common urea (at comparable total N rates) increased leaf area index (LAI), relative chlorophyll content index (CCI) and PARi , leading to higher grain yield and NUE (increased 24.4% and 25.3% in early and late rice, respectively) compared to local farming practice. Structural equation model (SEM) analysis showed that differences in N application, between CR and common urea, directly accounted for differences observed in soil nutrient, PARi and NUE rather than yield components. Additionally, compared to traditional yield determinants, LAI and PARi (between booting and filling stage) are capable of predicting and explaining grain yield by 0.69 and 0.92 of R2 in early and late rice, respectively. CONCLUSION: Leaf morpho-physiological traits are important for developing N management practices to increase NUE and improve food security for paddy agriculture in southern China. © 2022 Society of Chemical Industry.

High-precision distributed detection of rail defects by tracking the acoustic propagation waves.

Nowadays, early defect detection plays a significant role for the railway safety warning. However, the existing methods cannot satisfy the requirements of real-time and high-precision detection. Here, a high-precision, distributed and on-line method for detecting rail defect is proposed and demonstrated. When a train goes through defects, the instantaneous elastic waves will be excited by the wheel-rail interaction, which will further propagate along railway tracks bidirectionally. Through mounting the backscattering enhanced optical fiber on the railway as sensors, the fiber optic distributed acoustic sensing system can record the propagation trace precisely. Further, the acoustic propagation fitting method is applied onto the propagation data to detect and locate defects along the long-distance railway. Especially, the dual-frequency joint-processing algorithm is proposed to improve the location accuracy. The field test proves that multiple defects along the railway can be successfully identified and located with a standard deviation of 0.314m. To the best of our knowledge, this work is the first report of distributed rail defect detection, which will bring a breakthrough for high-precision structural damage detection in the infrastructures such as the railway, pipeline and tunnel.

Therapeutic potential of targeting LSD1/ KDM1A in cancers.

LSD1 was the first histone demethylase identified by Professor Shi Yang and his team members in 2004. LSD1 employs FAD as its cofactor, which catalyzes the demethylation of H3K4 and H3K9. It is aberrantly overexpressed in different types of cancers and is associated with the growth, invasion, and metastasis of cancer cells. The knockout or inhibition of LSD1 could effectively suppress tumor development, and thus, it has become an attractive molecular target for cancer therapy. Moreover, many LSD1 inhibitors have been developed in preclinical and clinical trials to treat solid tumors and hematological malignancy. This study made an extensive review of the research obtained from the literature retrieval of electronic databases, such as PubMed, Web of Science, RCSB PDB, ClinicalTrials.gov, and EU clinical trials register. This review summarizes recent studies on the advances of LSD1 inhibitors in the literature, covering January 2015 to June 2021. It focuses on the function of LSD1 in tumor cells, summarizes the crystal structures of Homo sapiens LSD1, reviews the structural characteristics of LSD1 inhibitors, compares the screening methods of LSD1 inhibitors, and proposes guidelines for the future exploitation of LSD1 inhibitors.

Design, synthesis, and biological evaluation of potent FAK-degrading PROTACs.

FAK mediated tumour cell migration, invasion, survival, proliferation and regulation of tumour stem cells through its kinase-dependent enzymatic functions and kinase-independent scaffolding functions. At present, the development of FAK PROTACs has become one of the hotspots in current pharmaceutical research to solve above problems. Herein, we designed and synthesised a series of FAK-targeting PROTACs consisted of PF-562271 derivative 1 and Pomalidomide. All compounds showed significant in vitro FAK kinase inhibitory activity, the IC50 value of the optimised PROTAC A13 was 26.4 nM. Further, A13 exhibited optimal protein degradation (85% degradation at 10 nM). Meantime, compared with PF-562271, PROTAC A13 exhibited better antiproliferative activity and anti-invasion ability in A549 cells. More, A13 had excellent plasma stability with T1/2 >194.8 min. There are various signs that PROTAC A13 could be useful as expand tool for studying functions of FAK in biological system and as potential therapeutic agents.

Design, synthesis, and biological evaluation of coumarin analogs as novel LSD1 inhibitors.

The abnormal expression of lysine-specific histone demethylase 1 (LSD1) is associated with different cancer types, and it is increasingly recognized as a potential therapeutic target in oncology. Here, utilizing core hopping and conformational restriction strategies, we designed and synthesized a series of coumarin analogs that were shown to be potent LSD1 inhibitors in the enzyme assay. Furthermore, several potent compounds were selected to evaluate their antiproliferative activity on A549 cells and MGC-803 cells with high expression of LSD1. Among them, YX10 showed an anticlonogenic effect on A549 cells and MGC-803 cells, with IC50 values of 1.52 ± 0.16 and 0.98 ± 0.18 µM, respectively. Modeling suggested that the inhibitors would bind to the active site of the protein located around the key residues of Asp555 and Lys661. Meanwhile, a preliminary druggability evaluation showed that compound YX10 showed favorable liver microsomal and moderate plasma stability and weak inhibitory activity against cytochrome P450 isoforms at 10 µM. All the results indicated that compound YX10 could represent a promising lead compound for further development.

Design, synthesis, and evaluation of novel 3,4-isoxazolediamide derivatives for the combination treatment of azole-resistant candidiasis.

Invasive fungal infections are emerging as serious infectious diseases worldwide. Due to the frequent emergence of resistance, the cure for invasive fungal infections is often unachievable. The molecular chaperone Hsp90 provides a promising target because it supports survival, virulence, and drug resistance in a variety of pathogens. Herein, we report on the structural optimization and structure-activity relationship studies of 3,4-isoxazolediamide analogs. As a new class of fungal Hsp90 inhibitor, compound B25 was found to have good synergistic effects with fluconazole and to avoid potential mammalian toxicity. It also showed remarkable metabolic stability in vitro. Collectively, B25 could be a promising lead compound for drug discovery targeting fungal Hsp90 and deserves further investigation.

Design, synthesis, and biological evaluation of 5-aminotetrahydroquinoline-based LSD1 inhibitors acting on Asp375.

The abnormal expression of lysine-specific histone demethylase 1 (LSD1) is associated with different cancer types, and LSD1 inhibitory activity seems to have high therapeutic potential in cancer treatment. Here, we report the design, synthesis, and biochemical evaluation of novel 5-aminotetrahydroquinoline-based LSD1 inhibitors. Among them, compounds A6, A8, B1-B5, and C4 showed preferable inhibitory effects on LSD1, with IC50 = 0.19-0.82 µM. Several potent compounds were selected to evaluate their antiproliferative activity on A549 cells and MCF-7 cells with a high expression of LSD1. The potential binding modes of the compounds were revealed through molecular docking to rationalize the potency of compounds toward LSD1. Our data recognized that the 5-aminotetrahydroquinoline scaffold may serve as a starting point for developing potent LSD1 inhibitors for cancer therapy.

Producing more grain with lower environmental costs.

Agriculture faces great challenges to ensure global food security by increasing yields while reducing environmental costs. Here we address this challenge by conducting a total of 153 site-year field experiments covering the main agro-ecological areas for rice, wheat and maize production in China. A set of integrated soil-crop system management practices based on a modern understanding of crop ecophysiology and soil biogeochemistry increases average yields for rice, wheat and maize from 7.2 million grams per hectare (Mg ha(-1)), 7.2 Mg ha(-1) and 10.5 Mg ha(-1) to 8.5 Mg ha(-1), 8.9 Mg ha(-1) and 14.2 Mg ha(-1), respectively, without any increase in nitrogen fertilizer. Model simulation and life-cycle assessment show that reactive nitrogen losses and greenhouse gas emissions are reduced substantially by integrated soil-crop system management. If farmers in China could achieve average grain yields equivalent to 80% of this treatment by 2030, over the same planting area as in 2012, total production of rice, wheat and maize in China would be more than enough to meet the demand for direct human consumption and a substantially increased demand for animal feed, while decreasing the environmental costs of intensive agriculture.

Discovery of 2,4-diaminopyrimidine derivatives targeting p21-activated kinase 4: Biological evaluation and docking studies.

In this study, novel 2,4-diaminopyrimidine derivatives targeting p21-activated kinase 4 (PAK4) were discovered and evaluated for their biological activity against PAK4. Among the derivatives studied, promising compounds A2, B6, and B8 displayed the highest inhibitory activities against PAK4 (IC50 = 18.4, 5.9, and 20.4 nM, respectively). From the cellular assay, compound B6 exhibited the highest potency with an IC50 value of 2.533 µM against A549 cells. Some compounds were selected for computational ADME (absorption, distribution, metabolism, and elimination) properties and molecular docking studies against PAK4. The detailed structure-activity relationship based on the biochemical activities and molecular docking studies were explored. According to the docking studies, compound B6 had the lowest docking score (docking energy: -7.593 kcal/mol). The molecular docking simulation indicated the binding mode between compound B6 and PAK4. All these results suggest compound B6 as a useful candidate for the development of a PAK4 inhibitor.

Integrated systematic approach increase greenhouse tomato yield and reduce environmental losses.

High input - high output greenhouse vegetable systems are responsible for nutrient surpluses and environmental losses. Integrated strategies that improve soil, crop and nutrient management are needed to ensure more sustainable production systems. We conducted a two-year field experiment to evaluate the potential of integrated soil-crop system management (ISSM) practices to improve the productivity and environmental outcomes from an intensive greenhouse tomato production system in the Yangtze River Basin, China. Four treatments were tested: i) farmers' practice (FP); ii) soil remediation (SR), where lime nitrogen with compost addition was the only management strategy; iii) a treatment that combined soil remediation with optimized crop planting density (SRCO), which increased planting density for improving crop yield; and iv) integrated soil-crop system management (ISSM), as a systematic integrated approach, which included the combined optimization of soil remediation, crop optimization, and nutrient management. In the integrated soil-crop system management treatment, nutrient management was optimized through adoption of the most appropriate type (formula) of fertilizer for the crop, rate and application timing of synthetic fertilizer, and by substituting poultry manure with compost. Our results indicated that the fruit yield of the integrated soil-crop system management treatment was 104 t ha-1, 13.4%-37.3% higher than that of the other three treatments. The mean reactive nitrogen loss (81.1 kg N ha-1) and the greenhouse gas emissions (6495 kg CO2-eq ha-1) in the farmers' practice treatment were much higher than in the other three treatments (reactive nitrogen loss: 47.9-54.3 kg N ha-1; and greenhouse gas emissions: 4926-5468 kg CO2-eq ha-1, respectively). The mean nitrogen and carbon footprints of the integrated soil-crop system management treatment were significantly lower than those of other treatments, as a result of both the lower fertilizer nitrogen use and the greater yield. This study indicates that integrated soil-crop system management could produce greater yields and increase net profit with reduced nitrogen inputs, whilst reducing the environmental cost associated with conventional farmers' practice in plastic-greenhouse vegetable production systems.

Can microplastics variability drive the colonization dynamics of periphytic protozoan fauna in marine environments?

In recent years, microplastics have become a global environmental hot topic of concern. To explore the effects of different concentrations of microplastics on colonization dynamics of periphytic protozoan fauna, a 21-day study was conducted in temperature-controlled circulation systems. Periphytic protozoan communities were used as test organisms and exposed to five concentrations of MPs: 0, 1, 5, 25, and 125 mg l-1, identification and enumeration were conducted on days 3, 5, 7, 10, 14 and 21. The results showed that the colonization dynamics were driven by MPs and significantly shifted at concentrations over 5 mg l-1. However, a notable decline in maximum species richness and abundance was observed in the high concentrations of microplastic, along with significant deviations in colonization patterns from the control group (0 mg l-1). Therefore, it is suggested that the colonization dynamics of periphytic protozoa can serve as a bioindicator for assessing microplastic concentrations in marine environments.

Low-light enhancement method with dual branch feature fusion and learnable regularized attention.

Restricted by the lighting conditions, the images captured at night tend to suffer from color aberration, noise, and other unfavorable factors, making it difficult for subsequent vision-based applications. To solve this problem, we propose a two-stage size-controllable low-light enhancement method, named Dual Fusion Enhancement Net (DFEN). The whole algorithm is built on a double U-Net structure, implementing brightness adjustment and detail revision respectively. A dual branch feature fusion module is adopted to enhance its ability of feature extraction and aggregation. We also design a learnable regularized attention module to balance the enhancement effect on different regions. Besides, we introduce a cosine training strategy to smooth the transition of the training target from the brightness adjustment stage to the detail revision stage during the training process. The proposed DFEN is tested on several low-light datasets, and the experimental results demonstrate that the algorithm achieves superior enhancement results with the similar parameters. It is worth noting that the lightest DFEN model reaches 11 FPS for image size of 1224×1024 in an RTX 3090 GPU.

Impact of biological manure substitution on grain yield, nitrogen recovery efficiency, and soil biochemical properties.

Fertilization plays a crucial role in ensuring global food security and ecological balance. This study investigated the impact of substituting innovative biological manure for chemical fertilization on rice (Oryza sativa L) productivity and soil biochemical properties based on a three-year experiment. Our results suggested rice yield and straw weight were increased under manure addition treatment. Specifically, 70% of total nitrogen (N) fertilizer substituted by biological manure derived from straw, animal waste and microbiome, led to a substantial 13.6% increase in rice yield and a remarkable 34.2% boost in straw weight. In comparison to the conventional local farmer practice of applying 165 kg N ha-1, adopting 70% of total N plus biological manure demonstrated superior outcomes, particularly in enhancing yield components and spike morphology. Fertilization treatments led to elevated levels of soil microbial biomass carbon and N. However, a nuanced comparison with local practices indicated that applying biological manure alongside urea resulted in a slight reduction in N content in vegetative and economic organs, along with decreases of 10.4%, 11.2%, and 6.1% in N recovery efficiency (NRE), respectively. Prudent N management through the judicious application of partial biological manure fertilizer in rice systems could be imperative for sustaining productivity and soil fertility in southern China.

Discovery of novel 3-(1H-pyrazol-4-yl)-1H-indazole derivatives as potent type II TRK inhibitors against acquired resistance.

Tropomyosin receptor kinase (TRK) is a promising target for treating NTRK fusion cancers. The solvent front and xDFG mutations induced by larotrectinib and entrectinib result in acquired resistance in advanced-stage patients. In this study, we report a highly potent and selective type II TRK inhibitor, 40l, developed using a structure-based design strategy. Compound 40l significantly suppressed Km-12, Ba/F3-TRKAG595R, and Ba/F3-TRKAG667C cell proliferation. In biochemical and cellular assays, 40l showed better inhibitory activity against TRKAG667C than that by the positive control, selitrectinib. Additionally, it induced apoptosis of Ba/F3-TRKAG595R and Ba/F3-TRKAG667C cells in a dose-dependent manner. Furthermore, 40l showed good selectivity for a panel of 41 kinases. In vitro assays indicated that 40l possessed outstanding plasma stability and moderate liver microsomal stability. Based on the above results, compound 40l could be further optimized to overcome the solvent front and xDFG TRK mutations.