Summary
In eukaryotes, mechanisms such as alternative splicing (AS) and alternative translation initiation (ATI) contribute to organismal protein diversity. Specifically, splicing factors play ...crucial roles in responses to environment and development cues; however, the underlying mechanisms are not well investigated in plants. Here, we report the parallel employment of short‐read RNA sequencing, single molecule long‐read sequencing and proteomic identification to unravel AS isoforms and previously unannotated proteins in response to abscisic acid (ABA) treatment. Combining the data from the two sequencing methods, approximately 83.4% of intron‐containing genes were alternatively spliced. Two AS types, which are referred to as alternative first exon (AFE) and alternative last exon (ALE), were more abundant than intron retention (IR); however, by contrast to AS events detected under normal conditions, differentially expressed AS isoforms were more likely to be translated. ABA extensively affects the AS pattern, indicated by the increasing number of non‐conventional splicing sites. This work also identified thousands of unannotated peptides and proteins by ATI based on mass spectrometry and a virtual peptide library deduced from both strands of coding regions within the Arabidopsis genome. The results enhance our understanding of AS and alternative translation mechanisms under normal conditions, and in response to ABA treatment.
Significance Statement
In this study, a customized analytical pipeline was developed to study transcriptional and translational changes during the abscisic acid response in plants. Using single molecule long‐read sequencing and short‐read RNA sequencing, we identified numerous alternative spliced (AS) transcripts in Arabidopsis and characterized two new AS types. Proteomic identification indicates differentially expressed AS events were more likely to undergo protein translation. The entire workflow is applicable for other plant species.
To our knowledge, no randomised study has compared postmastectomy hypofractionated radiotherapy with conventional fractionated radiotherapy in patients with breast cancer. This study aimed to ...determine whether a 3-week schedule of postmastectomy hypofractionated radiotherapy is as efficacious and safe as a 5-week schedule of conventional fractionated radiotherapy.
This randomised, non-inferiority, open-label, phase 3 study was done in a single academic hospital in China. Patients aged 18–75 years who had undergone mastectomy and had at least four positive axillary lymph nodes or primary tumour stage T3–4 disease were eligible to participate. Patients were randomly assigned (1:1) according to a computer-generated central randomisation schedule, without stratification, to receive chest wall and nodal irradiation at a dose of 50 Gy in 25 fractions over 5 weeks (conventional fractionated radiotherapy) or 43·5 Gy in 15 fractions over 3 weeks (hypofractionated radiotherapy). The modified intention-to-treat population (including all eligible patients who underwent randomisation but excluding those who were considered ineligible or withdrew consent after randomisation) was used in primary and safety analyses. The primary endpoint was 5-year locoregional recurrence, and a 5% margin was used to establish non-inferiority (equivalent to a hazard ratio <1·883). This trial is registered at ClinicalTrials.gov, number NCT00793962.
Between June 12, 2008, and June 16, 2016, 820 patients were enrolled and randomly assigned to the conventional fractionated radiotherapy group (n=414) or hypofractionated radiotherapy group (n=406). 409 participants in the conventional fractionated radiotherapy group and 401 participants in the hypofractionated radiotherapy group were included in the modified intention-to-treat analyses. At a median follow-up of 58·5 months (IQR 39·2–81·8), 60 (7%) patients had developed locoregional recurrence (31 patients in the hypofractionated radiotherapy group and 29 in the conventional fractionated radiotherapy group); the 5-year cumulative incidence of locoregional recurrence was 8·3% (90% CI 5·8–10·7) in the hypofractionated radiotherapy group and 8·1% (90% CI 5·4–10·6) in the conventional fractionated radiotherapy group (absolute difference 0·2%, 90% CI −3·0 to 2·6; hazard ratio 1·10, 90% CI 0·72 to 1·69; p<0·0001 for non-inferiority). There were no significant differences between the groups in acute and late toxicities, except that fewer patients in the hypofractionated radiotherapy group had grade 3 acute skin toxicity than in the conventional fractionated radiotherapy group (14 3% of 401 patients vs 32 8% of 409 patients; p<0·0001).
Postmastectomy hypofractionated radiotherapy was non-inferior to and had similar toxicities to conventional fractionated radiotherapy in patients with high-risk breast cancer. Hypofractionated radiotherapy could provide more convenient treatment and allow providers to treat more patients.
National Key Projects of Research and Development of China; the Chinese Academy of Medical Science Innovation Fund for Medical Sciences; and Beijing Marathon of Hope, Cancer Foundation of China.
Rigorous risk assessment of chemicals in food and feed is essential to address the growing worldwide concerns about food safety. High-quality toxicological data on food-relevant chemicals are ...fundamental for risk modeling and assessment in the food safety area. The organization and analysis of substantial toxicity information can positively support decision-making by providing insight into toxicity trends. However, it remains challenging to systematically obtain fragmented toxicity data, and related toxicological resources are required to meet the current demands. In this study, we collected 221,439 experimental toxicity records for 5,657 food-relevant chemicals identified from extensive databases and literature, along with their information on chemical identification, physicochemical properties, environmental fates, and biological targets. Based on the aggregated data, a freely available web-based databank, Food-Relevant Available Chemicals Toxicology Databank (FRAC-TD) is presented, which supports multiple browsing ways and search criterions. Applying FRAC-TD for data-driven analysis, we revealed the underlying toxicity profiles of food-relevant chemicals in humans, mammals, and other species in the food chain. Expectantly, FRAC-TD could positively facilitate toxicological studies, toxicity prediction, and risk assessments in the food industry.
Protoporphyrinogen IX oxidase (PPO/Protox, E.C. 1.3.3.4) is recognized as one of the most important targets for herbicide discovery. In this study, we report our ongoing research efforts toward the ...discovery of novel PPO inhibitors. Specifically, we identified a highly potent new compound series containing a pyrimidinedione moiety and bearing a versatile building block-benzoxazinone scaffold. Systematic bioassays resulted in the discovery of compound 7af, ethyl 4-(7-fluoro-6-(3-methyl-2,6-dioxo-4-(trifluoromethyl)-3,6-dihydropyrimidin-1(2H)-yl)-3-oxo-2,3-dihydro-4H-benzob1,4oxazin-4-yl)butanoate, which exhibited broad-spectrum and excellent herbicidal activity at the dosage of 37.5 g a.i./ha through postemergence application. The inhibition constant (K i) value of 7af to Nicotiana tabacum PPO (NtPPO) was 14 nM, while to human PPO (hPPO), it was 44.8 μM, indicating a selective factor of 3200, making it the most selective PPO inhibitor to date. Moreover, molecular simulations further demonstrated the selectivity and the binding mechanism of 7af to NtPPO and hPPO. This study not only identifies a candidate that showed excellent in vivo bioactivity and high safety toward humans but also provides a paradigm for discovering PPO inhibitors with improved performance through molecular simulation and structure-guided optimization.
4-Hydroxyphenylpyruvate dioxygenase (HPPD) is a functional protein existing in almost all aerobic organisms. In the field of agricultural chemicals, HPPD is acknowledged to be one of the crucial ...targets for herbicides at present due to its unique bio-function in plants. In the Auto Core Fragment in silico Screening (ACFIS) web server, a potential HPPD inhibitor featuring 1,2,3-benzotriazine-4-one was screened out via a pharmacophore-linked fragment virtual screening (PFVS) method. Molecular simulation studies drove the process of “hit-to-lead” optimization, and a family of 1,2,3-benzotriazine-4-one derivatives was synthesized. Consequently, 6-(2-hydroxy-6-oxocyclohex-1-ene-1-carbonyl)-5-methyl-3-(2-methylbenzyl)benzod1,2,3triazin-4(3H)-one (15bu) was identified to be the best HPPD inhibitor (IC50 = 36 nM) among the 1,2,3-benzotriazine-4-one derivatives, which had over 8-fold improvement of enzyme inhibition compared with the positive control mesotrione (IC50 = 289 nM). Crystallography information for the AtHPPD–15bu complex revealed several important interactions of the ligand bound upon the target protein, i.e., the bidentate chelating interaction of the triketone motif with the metal ion of AtHPPD, a tight π–π stacking interaction consisting of the1,2,3-benzotriazine-4-one moiety and two benzene rings of Phe-424 and Phe-381, and the polydirectional hydrophobic contacts consisting of the ortho-CH3-benzyl group of the core scaffold and some hydrophobic residues. Furthermore, compound 15bu displayed 100% inhibition against the five species of target weeds at the tested dosage, which was comparable to the weed control of mesotrione. Collectively, the fused 1,2,3-benzotriazine-4-one-triketone hybrid is a promising chemical tool for the development of more potent HPPD inhibitors and provides a valuable lead compound 15bu for herbicide innovation.
Abstract
Protein post-translational modifications (PTM) play vital roles in cellular regulation, modulating functions by driving changes in protein structure and dynamics. Exploring comprehensively ...the influence of PTM on conformational dynamics can facilitate the understanding of the related biological function and molecular mechanism. Currently, a series of excellent computation tools have been designed to analyze the time-dependent structural properties of proteins. However, the protocol aimed to explore conformational dynamics of post-translational modified protein is still a blank. To fill this gap, we present PTMdyna to visually predict the conformational dynamics differences between unmodified and modified proteins, thus indicating the influence of specific PTM. PTMdyna exhibits an AUC of 0.884 tested on 220 protein–protein complex structures. The case of heterochromatin protein 1α complexed with lysine 9-methylated histone H3, which is critical for genomic stability and cell differentiation, was used to demonstrate its applicability. PTMdyna provides a reliable platform to predict the influence of PTM on protein dynamics, making it easier to interpret PTM functionality at the structure level. The web server is freely available at http://ccbportal.com/PTMdyna.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Protein–protein interactions (PPIs) have vital roles in almost all cellular processes, although they are regarded as ‘undruggable’ therapeutic targets because of their large, flat, featureless ...interfaces.Fragment-based drug discovery (FBDD) shows advantages for the design of modulators to inhibit or stabilize PPIs, with two drugs approved and more than ten compounds in clinical trials.Fragments tend to bind at ‘hot spots’ of PPI interfaces, and the locations of different ‘hot spots’ provide directions for fragment evolution to discover more potent PPI modulators.Fragment hits with new mode of actions, including covalent fragments and fragments binding at allosteric sites, have become important sources of PPI modulators.Emerging technologies, such as cryo-electron microscopy, covalent tethering, and artificial intelligence, are accelerating the application of FBDD targeting PPIs.
Protein–protein interactions (PPIs) have important roles in various cellular processes, but are commonly described as ‘undruggable’ therapeutic targets due to their large, flat, featureless interfaces. Fragment-based drug discovery (FBDD) has achieved great success in modulating PPIs, with more than ten compounds in clinical trials. Here, we highlight the progress of FBDD in modulating PPIs for therapeutic development. Targeting hot spots that have essential roles in both fragment binding and PPIs provides a shortcut for the development of PPI modulators via FBDD. We highlight successful cases of cracking the ‘undruggable’ problems of PPIs using fragment-based approaches. We also introduce new technologies and future trends. Thus, we hope that this review will provide useful guidance for drug discovery targeting PPIs.
It has been reported that cadherin 6 (CDH6) upregulation is associated with enhanced epithelial-to-mesenchymal transition (EMT) in several types of solid tumor cells. The current study aimed to ...explore the effect of CDH6 on the migration and invasion of stomach adenocarcinoma (STAD) cells, the transcription factors involved in CDH6 dysregulation and their effect on mitochondrial fission. Bioinformatics analysis was performed using data extracted from the Genotype-Tissue Expression Project, the Cancer Genome Atlas and Kaplan-Meier plotter. AGS and HGC27 cells were used to establish an in vitro STAD cell model. The results showed that higher CDH6 expression was associated with significantly shorter overall survival in patients with STAD. In addition, CDH6 overexpression promoted wound healing, enhanced the invasion ability of tumor cells and increased mitochondrial fission. Glioma-associated oncogene family zinc finger 2 (GLI2) could bind to the CDH6 promoter and activate its transcription. Fluorescent labeling also showed that GLI2 overexpression promoted mitochondrial fission. However, CDH6 silencing significantly reduced mitochondrial fragmentation. Besides, GLI2 overexpression notably upregulated phosphorylated-focal adhesion kinase and dynamin-related protein 1. However, the above effects were largely abrogated by CDH6 knockdown. In conclusion, the present study suggested that the novel GLI2/CDH6 axis could enhance the migration, invasion and mitochondrial fission of STAD cells.
•CDH6 upregulation was associated with poor prognosis of stomach adenocarcinoma.•CDH6 positively modulates migration, invasion and mitochondrial fission of STAD cells.•GLI2 binds to the CDH6 promoter and activates its transcription.•GLI2-CDH6 axis increases STAD cell migration, invasion and mitochondrial fission.
The enzymatic activities of Furin, Transmembrane serine proteinase 2 (TMPRSS2), Cathepsin L (CTSL), and Angiotensin‐converting enzyme 2 (ACE2) receptor binding are necessary for the entry of ...coronaviruses into host cells. Precise inhibition of these key proteases in ACE2+ lung cells during a viral infection cycle shall prevent viral Spike (S) protein activation and its fusion with a host cell membrane, consequently averting virus entry to the cells. In this study, dual‐drug‐combined (TMPRSS2 inhibitor Camostat and CTSL inhibitor E‐64d) nanocarriers (NCs) are constructed conjugated with an anti‐human ACE2 (hACE2) antibody and employ Red Blood Cell (RBC)‐hitchhiking, termed “Nanoengineered RBCs,” for targeting lung cells. The significant therapeutic efficacy of the dual‐drug‐loaded nanoengineered RBCs in pseudovirus‐infected K18‐hACE2 transgenic mice is reported. Notably, the modular nanoengineered RBCs (anti‐receptor antibody+NCs+RBCs) precisely target key proteases of host cells in the lungs to block the entry of Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), regardless of virus variations. These findings are anticipated to benefit the development of a series of novel and safe host‐cell‐protecting antiviral therapies.
Dual‐drug‐combined (Camostat and E‐64d) nanocarriers (NCs) are constructed, conjugated with an anti‐hACE2 antibody and employed RBC‐hitchhiking, termed “Nanoengineered RBCs”, for effectively blocking SARS‐CoV‐2 cell entry in the lungs, regardless of virus variations. The nano‐micro system is modular (anti‐receptor antibody + drug‐loaded NCs + RBCs), and the findings will benefit the development of a series of novel and safe host‐cell‐protecting antiviral therapies.
•First report of Li isotope data in the Cryogenian clastic sedimentary strata.•Negative Li isotopic excursion in the basal Cryogenian interglacial interval.•Low Li isotope values suggested enhanced ...silicate weathering intensity.•Extensive seawater euxinia was triggered by intense silicate weathering.
The Cryogenian Period comprised two episodes of global glaciation (Sturtian and Marinoan glaciations) separated by a non-glacial interval, which was characterized by early radiations of eukaryotic algae and putative metazoans. Geochemical data indicate that the non-glacial interval might be marked by a transient marine oxygenation, nevertheless oceanic redox conditions varied both in time and space. Further, the links between non-glacial climate and marine redox variations are not well constrained. Here we present high-resolution lithium isotope (δ7Li), Fe speciation and trace element (Mo and U) data for clastic sedimentary rocks from the Cryogenian interglacial Datangpo Formation, South China, in order to track the evolution of continental chemical silicate weathering and driving factors behind marine redox variability during the Cryogenian non-glacial interval. A significantly negative δ7Li excursion of ∼−5‰ is observed in the basal Datangpo Formation, suggesting a dramatic increase in chemical silicate weathering intensity in the aftermath of the Sturtian glaciation. Expansion and contraction of anoxic-sulfidic conditions, as demonstrated by Fe speciation and trace element (Mo and U) data, mirror changes in silicate weathering intensity. Our study provides evidence that greater nutrient and sulfate availability, due to high silicate weathering intensity associated with increased exposure of fresh rocks and a warm climate, facilitated the spread of euxinic waters over the continental margins of the otherwise ferruginous Cryogenian ocean.
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