Auto-inhibition and activation of a short Argonaute-associated TIR-APAZ defense system.

Short prokaryotic Ago accounts for most prokaryotic Argonaute proteins (pAgos) and is involved in defending bacteria against invading nucleic acids. Short pAgo associated with TIR-APAZ (SPARTA) has been shown to oligomerize and deplete NAD+ upon guide-mediated target DNA recognition. However, the molecular basis of SPARTA inhibition and activation remains unknown. In this study, we determined the cryogenic electron microscopy structures of Crenotalea thermophila SPARTA in its inhibited, transient and activated states. The SPARTA monomer is auto-inhibited by its acidic tail, which occupies the guide-target binding channel. Guide-mediated target binding expels this acidic tail and triggers substantial conformational changes to expose the Ago-Ago dimerization interface. As a result, SPARTA assembles into an active tetramer, where the four TIR domains are rearranged and packed to form NADase active sites. Together with biochemical evidence, our results provide a panoramic vision explaining SPARTA auto-inhibition and activation and expand understanding of pAgo-mediated bacterial defense systems.

GCPDFFNet: Small Object Detection for Rice Blast Recognition.

Early detection of rice blast disease is pivotal to ensure rice yield. We collected in situ images of rice blast and constructed a rice blast dataset based on variations in lesion shape, size, and color. Given that rice blast lesions are small and typically exhibit round, oval, and fusiform shapes, we proposed a small object detection model named GCPDFFNet (global context-based parallel differentiation feature fusion network) for rice blast recognition. The GCPDFFNet model has three global context feature extraction modules and two parallel differentiation feature fusion modules. The global context modules are employed to focus on the lesion areas; the parallel differentiation feature fusion modules are used to enhance the recognition effect of small-sized lesions. In addition, we proposed the SCYLLA normalized Wasserstein distance loss function, specifically designed to accelerate model convergence and improve the detection accuracy of rice blast disease. Comparative experiments were conducted on the rice blast dataset to evaluate the performance of the model. The proposed GCPDFFNet model outperformed the baseline network CenterNet, with a significant increase in mean average precision from 83.6 to 95.4% on the rice blast test set while maintaining a satisfactory frames per second drop from 147.9 to 122.1. Our results suggest that the GCPDFFNet model can accurately detect in situ rice blast disease while ensuring the inference speed meets the real-time requirements.

ELF4 contributes to esophageal squamous cell carcinoma growth and metastasis by augmenting cancer stemness via FUT9.

Esophageal squamous cell carcinoma (ESCC) commonly has aggressive properties and a poor prognosis. Investigating the molecular mechanisms underlying the progression of ESCC is crucial for developing effective therapeutic strategies. Here, by performing transcriptome sequencing in ESCC and adjacent normal tissues, we find that E74-like transcription factor 4 (ELF4) is the main upregulated transcription factor in ESCC. The results of the immunohistochemistry show that ELF4 is overexpressed in ESCC tissues and is significantly correlated with cancer staging and prognosis. Furthermore, we demonstrate that ELF4 could promote cancer cell proliferation, migration, invasion, and stemness by in vivo assays. Through RNA-seq and ChIP assays, we find that the stemness-related gene fucosyltransferase 9 ( FUT9) is transcriptionally activated by ELF4. Meanwhile, ELF4 is verified to affect ESCC cancer stemness by regulating FUT9 expression. Overall, we first discover that the transcription factor ELF4 is overexpressed in ESCC and can promote ESCC progression by transcriptionally upregulating the stemness-related gene FUT9.

Hsp90α promotes chemoresistance in pancreatic cancer by regulating Keap1-Nrf2 axis and inhibiting ferroptosis.

Chemoresistance is the primary reason for poor prognosis in patients with pancreatic cancer (PC). Recent studies have indicated that ferroptosis may improve chemoresistance, but the underlying mechanisms remain unclear. In this study, significant upregulation of heat shock protein 90α (Hsp90α) expression is detected in the peripheral blood and tissue samples of patients with chemoresistant PC. Further studies reveal that Hsp90α promotes the proliferation, migration, and invasion of a chemoresistant pancreatic cell line (Panc-1-gem) by suppressing ferroptosis. Hsp90α competitively binds to Kelch-like ECH-associated protein 1 (Keap1), liberating nuclear factor erythroid 2-related factor 2 (Nrf2) from Keap1 sequestration. Nrf2 subsequently translocates into the nucleus and activates the glutathione peroxidase 4 (GPX4) pathway, thereby suppressing ferroptosis. This process further worsens the chemoresistance of PC cells. This study provides valuable insight into potential molecular targets to overcome chemoresistance in PC. It sheds light on the intricate mechanisms linking Hsp90α and ferroptosis to chemoresistance in PC and provides a theoretical foundation for the development of novel therapeutic strategies.

Homeobox A7 promotes esophageal squamous cell carcinoma progression through C-C motif chemokine ligand 2-mediated tumor-associated macrophage recruitment.

Homeobox A7 (HOXA7) plays essential roles in multiple malignancies and was reported to be overexpressed in esophageal squamous cell carcinoma (ESCC). However, its functions in the ESCC tumor microenvironment remain to be explored. In this study, we showed that HOXA7 was overexpressed in ESCC among HOXA family members and correlated with tumor-associated macrophage (TAM) infiltration both in The Cancer Genome Atlas database and ESCC clinical samples. Moreover, transactivation of C-C motif chemokine ligand 2 (CCL2) by HOXA7 was identified (real-time quantitative PCR [RT-qPCR], western blot analysis, ELISA, and ChIP-qPCR), which was detected to drive chemotaxis and M2 polarization of macrophages both in vitro (Transwell assay) and in vivo (xenograft tumors models). In addition, CCL2 triggers macrophage expression of epidermal growth factor (EGF) (RT-qPCR and ELISA), which promotes tumor proliferation and metastasis by activating its receptor EGFR. In addition, EGF-induced ESCC cell proliferation and migration can be abrogated by HOXA7 knockdown (CCK-8 proliferation assay, EdU fluorescence, and Transwell assay). These results indicate a novel mechanistic role of HOXA7 in the cross-talk between ESCC and TAMs, which could be an underlying therapeutic target for ESCC.

X-ray crystallography study and optimization of novel benzothiophene analogs as potent selective estrogen receptor covalent antagonists (SERCAs) with improved potency and safety profiles.

Endocrine therapy (ET) is a well-validated strategy for estrogen receptor α positive (ERα + ) breast cancer therapy. Despite the clinical success of current standard of care (SoC), endocrine-resistance inevitably emerges and remains a significant medical challenge. Herein, we describe the structural optimization and evaluation of a new series of selective estrogen receptor covalent antagonists (SERCAs) based on benzothiophene scaffold. Among them, compounds 15b and 39d were identified as two highly potent covalent antagonists, which exhibits superior antiproliferation activity than positive controls against MCF-7 cells and shows high selectivity over ERα negative (ERα-) cells. More importantly, their mode of covalent engagement at Cys530 residue was accurately illustrated by a cocrystal structure of 15b-bound ERαY537S (PDB ID: 7WNV) and intact mass spectrometry, respectively. Further in vivo studies demonstrated potent antitumor activity in MCF-7 xenograft mouse model and an improved safety profile. Collectively, these compounds could be promising candidates for future development of the next generation SERCAs for endocrine-resistant ERα + breast cancer.

Effect of chemotherapy and different chemotherapy regimens on bone health among Chinese breast cancer women in different menstrual status: a self-control study.

PURPOSE: Although the therapy-related bone loss attracts increasing attention nowadays, the differences in chemotherapy-induced bone loss and bone metabolism indexes change among breast cancer (BC) women with different menstrual statuses or chemotherapy regimens are unknown. The aim of the study is to explore the effects of different regimens of chemotherapy on bone health. METHOD: The self-control study enrolled 118 initially diagnosed BC women without distant metastasis who underwent dual-energy X-ray absorptiometry (DXA) bone mineral density (BMD) screening and (or) bone metabolism index monitoring during chemotherapy at Chongqing Breast Cancer Center. Mann-Whitney U test, Cochran's Q test, and Wilcoxon sign rank test were performed. RESULTS: After chemotherapy, the BMD in the lumbar 1-4 and whole lumbar statistically decreased (- 1.8%/per 6 months), leading to a significantly increased proportion of osteoporosis (27.1% vs. 20.5%, P < 0.05), which were mainly seen in the premenopausal group (- 7.0%/per 6 months). Of the chemotherapeutic regimens of EC (epirubicin + cyclophosphamide), TC (docetaxel + cyclophosphamide), TEC (docetaxel + epirubicin + cyclophosphamide), and EC-T(H) [epirubicin + cyclophosphamide-docetaxel and/or trastuzumab], EC regimen had the least adverse impact on BMD, while the EC-TH regimen reduced BMD most (P < 0.05) inspite of the non-statistical difference between EC-T regimen, which was mainly seen in the postmenopausal group. Chemotherapy-induced amenorrhea (estradiol 94 pg/ml vs, 22 pg/ml; FSH 9.33 mIU/ml vs. 61.27 mIU/ml) was proved in premenopausal subgroup (P < 0.001). Except the postmenopausal population with calcium/VitD supplement, the albumin-adjusted calcium increased significantly (2.21 mmol/l vs. 2.33 mmol/l, P < 0.05) after chemotherapy. In postmenopausal group with calcium/VitD supplement, ß-CTX decreased significantly (0.56 ng/ml vs. 0.39 ng/ml, P < 0.05) and BMD were not affected by chemotherapy (P > 0. 05). In premenopausal group with calcium/VitD supplement, PTH decreased significantly (52.90 pg/ml vs. 28.80 pg/ml, P = 0. 008) and hip BMD increased after chemotherapy (0.845 g/m2 vs. 0.952 g/m2, P = 0. 006). As for both postmenopausal and premenopausal group without calcium/VitD supplement, there was a significant decrease in bone mass in hip and lumbar vertebrae after chemotherapy (0.831 g/m2 vs. 0.776 g/m2; 0.895 g/m2 vs. 0.870 g/m2, P < 0.05). CONCLUSION: Chemotherapy might induce lumbar vertebrae BMD loss and spine osteoporosis with regimen differences among Chinese BC patients. Calcium/VitD supplementation could improve bone turnover markers, bone metabolism indicators, and bone mineral density. Early interventions on bone health are needed for BC patients during chemotherapy.

Asiaticoside Mitigates Alzheimer's Disease Pathology by Attenuating Inflammation and Enhancing Synaptic Function.

Alzheimer's disease (AD) is a prevalent neurodegenerative disorder, hallmarked by the accumulation of amyloid-ß (Aß) plaques and neurofibrillary tangles. Due to the uncertainty of the pathogenesis of AD, strategies aimed at suppressing neuroinflammation and fostering synaptic repair are eagerly sought. Asiaticoside (AS), a natural triterpenoid derivative derived from Centella asiatica, is known for its anti-inflammatory, antioxidant, and wound-healing properties; however, its neuroprotective function in AD remains unclear. Our current study reveals that AS, when administered (40 mg/kg) in vivo, can mitigate cognitive dysfunction and attenuate neuroinflammation by inhibiting the activation of microglia and proinflammatory factors in Aß1-42-induced AD mice. Further mechanistic investigation suggests that AS may ameliorate cognitive impairment by inhibiting the activation of the p38 MAPK pathway and promoting synaptic repair. Our findings propose that AS could be a promising candidate for AD treatment, offering neuroinflammation inhibition and enhancement of synaptic function.

Population-specific cut-off points of fatty liver index: a study based on the National Health and Nutrition Examination Survey data.

BACKGROUND: Fatty liver index (FLI) is the most recognized blood biomarker for diagnosis of hepatic steatosis (HS), but lacks the reliable specific cut-off points (COPs). Therefore, we aim to investigate the population-specific COPs of FLI based on the results of liver ultrasound transient elastography (LUTE) and conventional ultrasonography in the National Health and Nutrition Examination Survey (NHANES). METHODS: 5948 participants who underwent LUTE from the NHANES 2017-2018 and 14,797 participants who underwent conventional ultrasonography from the Third NHANES (NHANES III) were recruited. FLI was calculated by using body mass index (BMI), waist circumference (WC), triglyceride, and gamma-glutamyl transferase, and its optimal COPs in a specific population (stratified by sex, BMI, and WC) were obtained from receiver operator characteristics (ROC) curve with ultrasonic-diagnosed HS as the reference standard. RESULTS: Based on LUTE in NHANES 2017-2018, the prevalence of HS and metabolic dysfunction-associated fatty liver disease (MAFLD) were 58.7% and 56.2%, respectively, and the optimal COP of FLI for HS diagnosis in the overall population was 45.60, with an area under ROC curve (AUROC) of 0.833 (0.822-0.844). Based on conventional ultrasonography in NHANES III, the prevalence of HS and MAFLD were 34.4% and 27. 9%, respectively, and the optimal COP of FLI for HS was 59.5, with an AUROC of 0.681 (0.671-0.691). With the increase of BMI and WC, the COPs increased gradually with significant differences between different groups. Compared with conventional ultrasonography, the COPs of FLI based on LUTE were much more precise, with higher diagnostic ability. The population-specific COPs of FLI stratified by gender, WC, and BMI were tabulated. CONCLUSION: In the United States, the incidences of HS and MAFLD were high, especially when assessed by LUTE. The FLI based on LUTE is well capable of predicting HS when stratified by gender, WC, and BMI.

ETV5 overexpression promotes progression of esophageal squamous cell carcinoma by upregulating SKA1 and TRPV2.

Esophageal squamous cell carcinoma (ESCC) is notorious for the rapid progression especially early tumor metastasis due to the unclear mechanism. Recently, ETV5 attracts much attention for its potential role as an oncogenic transcription factor involved in multiple cancers. However, no one reported the mechanism behind the association between ETV5 expression and esophageal squamous cell carcinoma progression. In this study, we found that ETV5 was upregulated in ESCC both from online database and our ESCC tissues and ETV5 was associated with tumor staging and prognosis. Knockdown of ETV5 or its downstream genes SKA1 and TRPV2 significantly suppress ESCC cells migration and invasion, respectively. Additionally, in vivo study showed knockdown of ETV5 inhibited tumor metastasis. Further experiments unveiled ETV5 could transcriptionally upregulate the expression of SKA1 and TRPV2 and further activate MMPs in ESCC progression. In conclusion, ETV5 was associated with ESCC tumor staging and ESCC prognosis clinically. ETV5 promoted metastasis of ESCC by activating MMPs through augmenting the transcription of SKA1 and TRPV2. ETV5 was likely to be a novel oncogene and therapeutic target in ESCC.

Magnoflorine from Coptis chinese has the potential to treat DNCB-induced Atopic dermatits by inhibiting apoptosis of keratinocyte.

AIMS: In Sheng Nong's herbal classic in China, Rhizoma coptidisa(RC) could be used to treat Atopic dermatitsb(AD), but its core ingredient(s) and mechanism remains unknown. The present study aimed to find out the ingredients against AD and expound its mechanisms. MATERIALS AND METHODS: Seven alkaloids were isolated from RC to compare the inhibition against HaCaT cells by MTT assays and apoptosis of cells stimulated with TNF-α/IFN-γ by flow cytometry. The effects of target alkaloids against AD were evaluated on DNCBc (2,4-dinitrochlorobenzene)-induced atopic dermatitis in mice. KEY FINDINGS: Seven alkaloids were isolated from RC successfully. The results from MTT and flow cytometry indicated that among these alkaloids, only magnoflorine d(MAG) had no obvious toxicity on cells, but could inhibit the apoptosis of the cells stimulated with TNF-α/IFN-γ. Further animal experiments confirmed that MAG significantly attenuated the AD-like symptom and inhibited the AD-induced increases in IgE/IL-4, as compared with control (P < 0.01). Moreover, MAG reduced the low Δψme(mitochondrial membrane potential) in HaCaT cells. The results of western blotting proved that MAG inhibited apoptosis of keratinocytes through decreasing the expressions of CTSBf (cathepsin B), Cyte Cg (cytochrome C), Bid and caspase-3/7/8/9. SIGNIFICANCE: Overall, MAG inhibited apoptosis by decreasing the expression of apoptotic pathway-related proteins, and laid a foundation for the study of AD mechanisms.

Correlation between microbes and colorectal cancer: tumor apoptosis is induced by sitosterols through promoting gut microbiota to produce short-chain fatty acids.

The diversity of the bacterial community in the gut is closely related to human health. Gut microbes accomplish multiple physiological and biochemical functions. Sitosterols are a series of phytochemicals that have multiple pharmacological activities and are used as cholesterol-lowering drugs in clinical practice. In this study, we investigated the roles of bacteria and short-chain fatty acids (SCFAs) to the anti-colorectal cancer (anti-CRC) effects of sitosterols in BALB/c nude mice. Sitosterols were administered orally and gut microbiota composition and intestinal SCFAs changes were analyzed. The correlation between gut microbiota, SCFAs, and tumor apoptosis was assessed by a series of in vivo and in vitro experiments. Tumor growth in the mice was inhibited by sitosterol-treatment. Mechanistic studies revealed that sitosterol-treatment reduced the expression of PI3K/Akt, promoted the activation of Bad, decreased Bcl-xl, and enhanced cyto-c release, leading to caspase-9 and caspase-3 activation, PARP cleavage, and apoptosis. 16S rDNA analysis revealed that the diversity of microbiota, particularly phyla Bacteroidetes and Firmicutes, reduced dramatically in the gut of tumor-bearing mice, whilst treatment with sitosterols reversed these changes. The levels of SCFAs in the fecal samples of sitosterol-treated mice increased, leading to cancer cell apoptosis in vitro. Moreover, tumor apoptosis was induced after mice received a daily dose of 2 × 108 CFU/0.2 mL Lactobacillus pentosus or 20 mM/0.2 mL SCFAs. Taken together, these results demonstrate that sitosterols maintain a diverse microbial environment and enrich the content of L. pentosus in the gut, leading to the production of beneficial metabolites including SCFAs that promote tumor apoptosis.

A Novel Tropoloisoquinoline Alkaloid, Neotatarine, from Acorus calamus L.

A novel tropoloisoquinoline alkaloid, neotatarine (1), was isolated from the 95% ethanol extract of the rhizome parts of Acorus calamus L. The chemical structure was unambiguously elucidated by spectroscopic and single-crystal X-ray diffraction analysis. Neotatarine (1) exhibited significantly inhibitory activity against Aß25 - 35 induced PC12 cell death with 2, 4 and 8 µm comparing with the assay control (P < 0.01).

[A new sesquiterpene from Acorus calamus (â ¡)].

A new quaiane-tgpe sesquiterpene was isolated from the 95% ethanol extract of the rhizomes of Acorus calamus by silica gel and sephadex LH-20 column chromatographic methods. Structure and absolute configuration of the sesquiterpene were elucidated by spectroscopic data and X-ray crystallographic analysis, and named as 1R,5R,7S-guaiane-4R,10R-diol-6-one.

The Effect of Minimally Invasive Hematoma Aspiration on the JNK Signal Transduction Pathway after Experimental Intracerebral Hemorrhage in Rats.

OBJECTIVE: To explore the effect of minimally invasive hematoma aspiration (MIHA) on the c-Jun NH2-terminal kinase (JNK) signal transduction pathway after intracerebral hemorrhage (ICH). METHODS: In this experiment, 300 adult male Wistar rats were randomly and averagely divided into sham-operated group, ICH group and MIHA group. In each group, 60 rats were used in the detection of indexes in this experiment, while the other 40 rats were used to replace rats which reached the exclusion criteria (accidental death or operation failure). In ICH group and MIHA group, ICH was induced by injection of 70 µL of autologous arterial blood into rat brain, while only the rats in MIHA group were treated by MIHA 6 h after ICH. Rats in sham-operated group were injected nothing into brains, and they were not treated either, like rats in ICH group. In each group, six rats were randomly selected to observe their Bederson's scales persistently (6, 24, 48, 72, 96, 120 h after ICH). According to the time they were sacrificed, the remaining rats in each group were divided into 3 subgroups (24, 72, 120 h). The change of brain water content (BWC) was measured by the wet weight to dry weight ratio method. The morphology of neurons in cortex was observed by the hematoxylin-eosin (HE) staining. The expressions of phospho-c-Jun NH2-terminal kinase (pJNK) and JNK in peri-hematomal brain tissue were determined by the immunohistochemistry (IHC) and Western blotting (WB). RESULTS: At all time points, compared with the ICH groups, the expression of pJNK decreased obviously in MIHA groups (p < 0.05), while their Bederson's scales and BWC declined, and neuron injury in the cortex was relieved. The expression level of JNK was not altered at different groups. The data obtained by IHC and WB indicated a high-level of consistency, which provided a certain dependability of the test results. CONCLUSION: The JNK signal transduction pathway could be activated after intracerebral hemorrhage, with the expressions of pJNK increasing. MIHA could relieve the histo-pathological damage of nerve cells, reducing brain edema and neurological deficits, and these neuroprotective effects might be associated with suppression of JNK signal transduction pathway.

[Effect and mechanism of Vav3 on the proliferation of human gastric cancer SGC7901 cells].

OBJECTIVE: The purpose of this study was to investigate the effect and mechanism of Vav3 gene on the proliferation of human gastric cancer cell line SGC7901. METHODS: The expressions of Vav3 proten in gastric cancer tissue, tumor-adjacent tissue, human gastric cancer cell line SGC7901 and gastric epithelial cell line GES-1 cells were tested by Western blot. Vav3-siRNA was transfected into the SGC7901 cells. The proliferation of SGC7901 cells in vitro was measured by MTT assay. Cell cycle of SGC7901 cells was determined by flow cytometry.The expressions of proliferation-related genes PCNA, p16, cyclin D1, Rb were determined by qPCR and Western blot assay. Orthotopic transplantation nude mouse models of gastric cancer were prepared, and the tumor growth and expressions of PCNA, P16, cyclin D1, and Rb proteins were examined. RESULTS: The relative expressions of Vav3 in the gastric cancer and peritumoral tissue were 0.910±0.242 and 0.243±0.045, respectively; the relative expressions of Vav3 in SGC7901 and GSE-1 cells were 0.925±0.127 and 0.277±0.038, respevtively (both P<0.05). The expression of Vav3 protein in SGC7901 cells was effectively inhibited by Vav3-siRNA. Proliferation of SGC7901 cells was inhibited by (83.43±10.17)% after 80 nmol/L Vav3-siRNA transfection (P<0.05). The ratio of SGC7901 cells in G0/G1 phase was increased, and in S phase decreased after Vav3-siRNA transfection (both P<0.05). The expressions of PCNA and cyclin D1 were decreased in cells after Vav3-siRNA transfection, and expressions of p16 and Rb were increased after Vav3-siRNA transfection (P<0.05 for all). The tumor growth in the Vav3-siRNA group was much slower than that in the other 2 control groups of nude mouse models. Compared with the two control groups, expressions of PCNA and cyclin D1 were significantly lower in the Vav3-siRNA group, while expressions of p16 and Rb were increased (P<0.05 for all). CONCLUSION: Vav3 can promote the proliferation of gastric cancer cells by regulating proliferation-related genes.

Extrachromosomal circular DNA (eccDNA): from carcinogenesis to drug resistance.

Extrachromosomal circular DNA (eccDNA) is a circular form of DNA that exists outside of the chromosome. Although it has only been a few decades since its discovery, in recent years, it has been found to have a close relationship with cancer, which has attracted widespread attention from researchers. Thus far, under the persistent research of researchers from all over the world, eccDNA has been found to play an important role in a variety of tumors, including breast cancer, lung cancer, ovarian cancer, etc. Herein, we review the sources of eccDNA, classifications, and the mechanisms responsible for their biogenesis. In addition, we introduce the relationship between eccDNA and various cancers and the role of eccDNA in the generation and evolution of cancer. Finally, we summarize the research significance and importance of eccDNA in cancer, and highlight new prospects for the application of eccDNA in the future detection and treatment of cancer.

Precise Gene Knock-In Tools with Minimized Risk of DSBs: A Trend for Gene Manipulation.

Gene knock-in refers to the insertion of exogenous functional genes into a target genome to achieve continuous expression. Currently, most knock-in tools are based on site-directed nucleases, which can induce double-strand breaks (DSBs) at the target, following which the designed donors carrying functional genes can be inserted via the endogenous gene repair pathway. The size of donor genes is limited by the characteristics of gene repair, and the DSBs induce risks like genotoxicity. New generation tools, such as prime editing, transposase, and integrase, can insert larger gene fragments while minimizing or eliminating the risk of DSBs, opening new avenues in the development of animal models and gene therapy. However, the elimination of off-target events and the production of delivery carriers with precise requirements remain challenging, restricting the application of the current knock-in treatments to mainly in vitro settings. Here, a comprehensive review of the knock-in tools that do not/minimally rely on DSBs and use other mechanisms is provided. Moreover, the challenges and recent advances of in vivo knock-in treatments in terms of the therapeutic process is discussed. Collectively, the new generation of DSBs-minimizing and large-fragment knock-in tools has revolutionized the field of gene editing, from basic research to clinical treatment.

Screening for Active Compounds of Acorus calamus against SARS-CoV-2 Viral Protease and Mechanism Prediction.

COVID-19, caused by SARS-CoV-2, has emerged as the most destructive emerging infectious disease of the 21st century. Vaccination is an effective method to combat viral diseases. However, due to the constant mutation of the virus, new variants may weaken the efficacy of vaccines. In the current field of new coronavirus research, viral protease inhibitors have emerged as a highly regarded therapeutic strategy. Nevertheless, existing viral protease inhibitors do not fully meet the therapeutic needs. Therefore, this paper turned to traditional Chinese medicine to explore new active compounds. This study focused on 24 isolated compounds from Acorus calamus L. and identified 8 active components that exhibited significant inhibitory effects on SARS-CoV-2 PLpro. Among these, the compound 1R,5R,7S-guaiane-4R,10R-diol-6-one demonstrated the best inhibitory activity with IC50 values of 0.386 ± 0.118 µM. Additionally, menecubebane B and neo-acorane A exhibited inhibitory activity against both Mpro and PLpro proteases, indicating their potential as dual-target inhibitors. The molecular docking results confirmed the stable conformations of these compounds with the key targets and their good activity. ADMET and Lipinski's rule analyses revealed that all the small molecule ligands possessed excellent oral absorption properties. This study provides an experimental foundation for the discovery of promising antiviral lead compounds.

Mechanistic insights into lanthipeptide modification by a distinct subclass of LanKC enzyme that forms dimers.

Naturally occurring lanthipeptides, peptides post-translationally modified by various enzymes, hold significant promise as antibiotics. Despite extensive biochemical and structural studies, the events preceding peptide modification remain poorly understood. Here, we identify a distinct subclass of lanthionine synthetase KC (LanKC) enzymes with distinct structural and functional characteristics. We show that PneKC, a member of this subclass, forms a dimer and possesses GTPase activity. Through three cryo-EM structures of PneKC, we illustrate different stages of peptide PneA binding, from initial recognition to full binding. Our structures show the kinase domain complexed with the PneA core peptide and GTPγS, a phosphate-bound lyase domain, and an unconventional cyclase domain. The leader peptide of PneA interact with a gate loop, transitioning from an extended to a helical conformation. We identify a dimerization hot spot and propose a "negative cooperativity" mechanism toggling the enzyme between tense and relaxed conformation. Additionally, we identify an important salt bridge in the cyclase domain, differing from those in in conventional cyclase domains. These residues are highly conserved in the LanKC subclass and are part of two signature motifs. These results unveil potential differences in lanthipeptide modification enzymes assembly and deepen our understanding of allostery in these multifunctional enzymes.