Functional characterization of D-type cyclins involved in cell division in rice.

BACKGROUND: D-type cyclins (CYCD) regulate the cell cycle G1/S transition and are thus closely involved in cell cycle progression. However, little is known about their functions in rice. RESULTS: We identified 14 CYCD genes in the rice genome and confirmed the presence of characteristic cyclin domains in each. The expression of the OsCYCD genes in different tissues was investigated. Most OsCYCD genes were expressed at least in one of the analyzed tissues, with varying degrees of expression. Ten OsCYCD proteins could interact with both retinoblastoma-related protein (RBR) and A-type cyclin-dependent kinases (CDKA) forming holistic complexes, while OsCYCD3;1, OsCYCD6;1, and OsCYCD7;1 bound only one component, and OsCYCD4;2 bound to neither protein. Interestingly, all OsCYCD genes except OsCYCD7;1, were able to induce tobacco pavement cells to re-enter mitosis with different efficiencies. Transgenic rice plants overexpressing OsCYCD2;2, OsCYCD6;1, and OsCYCD7;1 (which induced cell division in tobacco with high-, low-, and zero-efficiency, respectively) were created. Higher levels of cell division were observed in both the stomatal lineage and epidermal cells of the OsCYCD2;2- and OsCYCD6;1-overexpressing plants, with lower levels seen in OsCYCD7;1-overexpressing plants. CONCLUSIONS: The distinct expression patterns and varying effects on the cell cycle suggest different functions for the various OsCYCD proteins. Our findings will enhance understanding of the CYCD family in rice and provide a preliminary foundation for the future functional verification of these genes.

PM2.5-derived exosomal long noncoding RNA PAET participates in childhood asthma by enhancing DNA damage via m6A-dependent OXPHOS regulation.

Fine particulate matter (PM2.5) is known to enhance DNA damage levels and is involved in respiratory diseases. Exosomes can carry noncoding RNAs, especially long noncoding RNAs (lncRNAs), as regulators of DNA damage, which participate in diseases. However, their role in PM2.5-induced childhood asthma remains unclear. We performed RNA-seq to profile aberrantly expressed exosomal lncRNAs derived from PM2.5-treated human bronchial epithelial (HBE) cell models. The role of exosomal lncRNAs in childhood asthma was determined in a case-control study. The intercellular communication mechanisms of exosomal lncRNA on DNA damage were determined in vitro. Exosomes secreted by PM2.5-treated HBE cells (PM2.5-Exos) could increase the DNA damage levels of recipient HBE cells and promote the expression levels of airway remodeling-related markers in sensitive human bronchial smooth muscle cells (HBSMCs). LncRNA PM2.5-associated exosomal transcript (PAET) was highly expressed in PM2.5-Exos and was associated with PM2.5 exposure in childhood asthma. Mechanistically, exosomal lncRNA PAET promoted methyltransferase-like 3 (METTL3) accumulation by increasing its stability, which stimulated N6-methyladenosine (m6A) modification of cytochrome c oxidase subunit 4I1 (COX4I1), and COX4I1 levels were decreased in a mechanism dependent on the m6A "reader" YTH domain family 3 (YTHDF3). COX4I1 deficiency subsequently disrupted oxidative phosphorylation (OXPHOS), resulting in attenuated adenosine triphosphate (ATP) production and accumulation of reactive oxygen species (ROS), which increased DNA damage levels. This comprehensive study extends the understanding of PM2.5-induced childhood asthma via DNA damage and identifies exosomal lncRNA PAET as a potential target for childhood asthma.

USP25 inhibits DNA damage by stabilizing BARD1 protein in a house dust mite-induced asthmatic model in vitro and in vivo.

House dust mites (HDM) can cause DNA double-strand breaks in the lungs of asthmatic patients. However, the molecular mechanisms driving DNA damage and repair in HDM-induced asthma are yet to be elucidated. Thus, in this study, HDM treatment was applied to BEAS-2B cells and mice to mimic the pathological process of asthma in vitro and in vivo, respectively. γ-H2AX foci and expression were measured by immunofluorescence staining and western blot, respectively. The levels of interleukin (IL)-4, IL-6, IL-13, and tumour necrosis factor α (TNFα) were measured using enzyme-linked immunoassay. The expression of USP25 and BARD1 was measured by reverse transcription quantitative PCR and western blot. Co-immunoprecipitation and ubiquitination assays were employed to detect the relationship between USP25 and BARD1. As per the results, it was found that the deubiquitylating enzyme USP25 repressed HDM-induced DNA damage and the production of proinflammatory cytokines, including TNF-α, IL-4, IL-8, and IL-13, in BEAS-2B cells; in contrast, the depletion of USP25 led to the opposite effects. USP25-mediated inhibition of DNA damage and inflammation was facilitated by the stabilizing protein BARD1, which is a tumor suppressor that principally functions by promoting DNA repair and replication in BEAS-2B cells. Furthermore, USP25 was found to robustly augment BARD1 protein abundance and prevent HDM-induced DNA damage and inflammation in vivo. Taken together, these results suggest a novel mechanism contributing to DNA damage and repair in HDM-induced asthma and that selectively modulating this pathway could lead to a novel therapeutic approach for controlling and managing asthma due to HDM exposure.

Deep learning applied to two-dimensional color Doppler flow imaging ultrasound images significantly improves diagnostic performance in the classification of breast masses: a multicenter study.

BACKGROUND: The current deep learning diagnosis of breast masses is mainly reflected by the diagnosis of benign and malignant lesions. In China, breast masses are divided into four categories according to the treatment method: inflammatory masses, adenosis, benign tumors, and malignant tumors. These categorizations are important for guiding clinical treatment. In this study, we aimed to develop a convolutional neural network (CNN) for classification of these four breast mass types using ultrasound (US) images. METHODS: Taking breast biopsy or pathological examinations as the reference standard, CNNs were used to establish models for the four-way classification of 3623 breast cancer patients from 13 centers. The patients were randomly divided into training and test groups (n = 1810 vs. n = 1813). Separate models were created for two-dimensional (2D) images only, 2D and color Doppler flow imaging (2D-CDFI), and 2D-CDFI and pulsed wave Doppler (2D-CDFI-PW) images. The performance of these three models was compared using sensitivity, specificity, area under receiver operating characteristic curve (AUC), positive (PPV) and negative predictive values (NPV), positive (LR+) and negative likelihood ratios (LR-), and the performance of the 2D model was further compared between masses of different sizes with above statistical indicators, between images from different hospitals with AUC, and with the performance of 37 radiologists. RESULTS: The accuracies of the 2D, 2D-CDFI, and 2D-CDFI-PW models on the test set were 87.9%, 89.2%, and 88.7%, respectively. The AUCs for classification of benign tumors, malignant tumors, inflammatory masses, and adenosis were 0.90, 0.91, 0.90, and 0.89, respectively (95% confidence intervals [CIs], 0.87-0.91, 0.89-0.92, 0.87-0.91, and 0.86-0.90). The 2D-CDFI model showed better accuracy (89.2%) on the test set than the 2D (87.9%) and 2D-CDFI-PW (88.7%) models. The 2D model showed accuracy of 81.7% on breast masses ≤1 cm and 82.3% on breast masses >1 cm; there was a significant difference between the two groups (P < 0.001). The accuracy of the CNN classifications for the test set (89.2%) was significantly higher than that of all the radiologists (30%). CONCLUSIONS: The CNN may have high accuracy for classification of US images of breast masses and perform significantly better than human radiologists. TRIAL REGISTRATION: Chictr.org, ChiCTR1900021375; http://www.chictr.org.cn/showproj.aspx?proj=33139.

Mitochondrial redox-driven mitofusin 2 S-glutathionylation promotes neuronal necroptosis via disrupting ER-mitochondria crosstalk in cadmium-induced neurotoxicity.

Reactive oxygen species (ROS)-mediated endoplasmic reticulum (ER) stress and mitochondrial dysfunction are known to affect the structural and functional damage in the neural system. Cadmium (Cd) is an environmental contaminant that is widely found in numerous environmental matrices and exhibits potential neurotoxic risk. However, it remains unclear how mitochondrial redox status induces, and whether Cd destabilizes, the ER-mitochondria crosstalk to have a toxic effect on the nervous system. Herein, in our present study, bioinformatics analysis revealed an important role of protein interaction and mitochondrial machinery in brain samples from Alzheimer's disease (AD) patients. Furthermore, we established a neurotoxicity model in vivo and in vitro induced by cadmium chloride (CdCl2). We demonstrated that CdCl2 exposure disrupts the balance in mitochondrial redox represented by enhanced mitochondrial ROS (mitoROS) levels, which enhance mitofusin 2 (Mfn2) S-glutathionylation and interrupt the mitochondria-associated ER membranes (MAMs) for crosstalk between the ER and mitochondria to induce neuronal necroptosis. Mechanistically, it was shown that CdCl2 exposure significantly enhances the mitochondria-associated degradation (MAD) of Mfn2 via S-glutathionylation, which inhibits Mfn2 localization to the MAMs and subsequently leads to the formation of the RIPK1-RIPK3-p-MLKL complex (a key component of the necrosome) at MAMs, to promote neuronal necroptosis. Furthermore, the glutaredoxin 1 (Grx1) catalyzed and Mfn2 overexpression restored S-glu-Mfn2, MAMs perturbation, necrosome formation, and necroptosis in neurons induced by CdCl2 exposure in vitro. Moreover, the intervention with antioxidants to reduce mitochondrial redox, such as N-acetyl-l-cysteine (NAC) and mitochondria-targeted antioxidant Mito-TEMPO, reduced the S-glutathionylation of Mfn2 involved in the antagonism of CdCl2-induced necroptosis and neurotoxicity in vivo and in vitro. Taken together, our results are the first time to demonstrate that S-glutathionylation of Mfn2 promotes neuronal necroptosis via disruption of ER-mitochondria crosstalk in CdCl2-induced neurotoxicity, providing the novel mechanistic insight into how hazardous chemical-induced adverse effects in various organs and tissues could be interpreted by intraorganellar pathways under the control of MAMs components in neurons.