765 results
Search Results
2. Comparative proteomic analysis of nuclear and cytoplasmic compartments in human cardiac progenitor cells.
- Author
-
Albericio G, Aguilar S, Torán JL, Yañez R, López JA, Vázquez J, Mora C, and Bernad A
- Subjects
- Cell Movement, Cell Proliferation, Cells, Cultured, Gene Expression Regulation, HMGA2 Protein genetics, HMGA2 Protein metabolism, Humans, Interleukin-1alpha genetics, Interleukin-1alpha metabolism, Oxidative Stress, RNA-Binding Proteins genetics, RNA-Binding Proteins metabolism, Receptor-Like Protein Tyrosine Phosphatases, Class 2 genetics, Receptor-Like Protein Tyrosine Phosphatases, Class 2 metabolism, Signal Transduction, Cell Nucleus metabolism, Cytoplasm metabolism, Fibroblasts metabolism, Mesenchymal Stem Cells metabolism, Myocytes, Cardiac metabolism, Proteome, Proteomics
- Abstract
Clinical trials evaluating cardiac progenitor cells (CPC) demonstrated feasibility and safety, but no clear functional benefits. Therefore a deeper understanding of CPC biology is warranted to inform strategies capable to enhance their therapeutic potential. Here we have defined, using a label-free proteomic approach, the differential cytoplasmic and nuclear compartments of human CPC (hCPC). Global analysis of cytoplasmic repertoire in hCPC suggested an important hypoxia response capacity and active collagen metabolism. In addition, comparative analysis of the nuclear protein compartment identified a significant regulation of a small number of proteins in hCPC versus human mesenchymal stem cells (hMSC). Two proteins significantly upregulated in the hCPC nuclear compartment, IL1A and IMP3, showed also a parallel increase in mRNA expression in hCPC versus hMSC, and were studied further. IL1A, subjected to an important post-transcriptional regulation, was demonstrated to act as a dual-function cytokine with a plausible role in apoptosis regulation. The knockdown of the mRNA binding protein (IMP3) did not negatively impact hCPC viability, but reduced their proliferation and migration capacity. Analysis of a panel of putative candidate genes identified HMGA2 and PTPRF as IMP3 targets in hCPC. Therefore, they are potentially involved in hCPC proliferation/migration regulation., (© 2022. The Author(s).)
- Published
- 2022
- Full Text
- View/download PDF
3. Structure Embedded Nucleus Classification for Histopathology Images.
- Author
-
Lou W, Wan X, Li G, Lou X, Li C, Gao F, and Li H
- Subjects
- Humans, Image Processing, Computer-Assisted methods, Histocytochemistry methods, Image Interpretation, Computer-Assisted methods, Cell Nucleus pathology, Neural Networks, Computer, Algorithms
- Abstract
Nuclei classification provides valuable information for histopathology image analysis. However, the large variations in the appearance of different nuclei types cause difficulties in identifying nuclei. Most neural network based methods are affected by the local receptive field of convolutions, and pay less attention to the spatial distribution of nuclei or the irregular contour shape of a nucleus. In this paper, we first propose a novel polygon-structure feature learning mechanism that transforms a nucleus contour into a sequence of points sampled in order, and employ a recurrent neural network that aggregates the sequential change in distance between key points to obtain learnable shape features. Next, we convert a histopathology image into a graph structure with nuclei as nodes, and build a graph neural network to embed the spatial distribution of nuclei into their representations. To capture the correlations between the categories of nuclei and their surrounding tissue patterns, we further introduce edge features that are defined as the background textures between adjacent nuclei. Lastly, we integrate both polygon and graph structure learning mechanisms into a whole framework that can extract intra and inter-nucleus structural characteristics for nuclei classification. Experimental results show that the proposed framework achieves significant improvements compared to the previous methods. Code and data are made available via https://github.com/lhaof/SENC.
- Published
- 2024
- Full Text
- View/download PDF
4. Different rates of pollen and seed gene flow cause branch‐length and geographic cytonuclear discordance within Asian butternuts
- Author
-
Bowen Zhang, Xin-Rui Lin, Lin-Lin Xu, Wei-Ning Bai, Rui-Min Yu, Kui Lin, Da-Yong Zhang, Nan Li, and CiiM, Zentrum für individualisierte Infektionsmedizin, Feodor-Lynen-Str.7, 30625 Hannover.
- Subjects
Gene Flow ,Nuclear gene ,Physiology ,Lineage (evolution) ,whole-genome resequencing ,introgression ,Introgression ,Plant Science ,whole‐genome resequencing ,Biology ,Genome ,Gene flow ,Coalescent theory ,Genome, Chloroplast ,Phylogeny ,Cell Nucleus ,Full Paper ,Research ,food and beverages ,Cline (biology) ,Full Papers ,branch-length discordance ,geographic discordance ,Evolutionary biology ,Seeds ,Pollen ,Biological dispersal ,branch‐length discordance ,gene flow - Abstract
Summary Topological cytonuclear discordance is commonly observed in plant phylogenetic and phylogeographic studies, yet few studies have attempted to detect two other forms of cytonuclear discordance (branch length and geographical) and to uncover the causes of the discordance.We used the whole nuclear and chloroplast genome data from 80 individual Asian butternuts to reveal the pattern and processes of cytonuclear discordance.Our findings indicate that the chloroplast genome had substantially deeper divergence (branch‐length discordance) and a steeper cline in the contact zone (geographic discordance) compared with the nuclear genome. After various hypothesis have been tested, the results suggest that incomplete lineage sorting, positive selection and cytonuclear incompatibility are probably insufficient to explain this pattern. However, isolation‐by‐distance analysis and gene flow estimation point to a much higher level of gene flow by pollen compared with by seeds, which may have slowed down lineage divergence and mediated wider contact for nuclear genome compared with the chloroplast genome.Altogether, this study highlights a critical role of sex‐biased dispersal in causing discordance between the nuclear and plastid genome of Asian butternuts. Given its ubiquity among plants, asymmetric gene flow should be given a high priority in future studies of cytonuclear discordance.
- Published
- 2021
5. The protective effects of Olmesartan against interleukin-29 (IL-29)-induced type 2 collagen degradation in human chondrocytes
- Author
-
Yunlong Liu, Junyi Liu, Yan Ma, Yongyong Zhang, Qiong Chen, Xin Yang, and Yanchun Shang
- Subjects
Cell Survival ,extracellular matrix ,Anti-Inflammatory Agents ,Tetrazoles ,Bioengineering ,Applied Microbiology and Biotechnology ,Models, Biological ,Cell Line ,Chondrocytes ,Transforming Growth Factor beta ,Osteoarthritis ,Olmesartan ,Humans ,Collagen Type II ,IL-29 ,Cell Proliferation ,Cell Nucleus ,Interleukins ,Imidazoles ,General Medicine ,Gene Expression Regulation ,Proteolysis ,TP248.13-248.65 ,Biotechnology ,Research Article ,Research Paper - Abstract
Osteoarthritis (OA) is a cartilage degenerative disease commonly observed in the elderly population and is pathologically characterized by the degradation of the cartilage extracellular matrix (ECM). Matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTSs) are critical enzymes involved in the degradation of ECM. Olmesartan is an inhibitor of the angiotensin II receptor developed for the treatment of hypertension, and recent studies show that it exerts anti-inflammatory effects in arthritis. The present study aimed to investigate the mechanism of the protective effect of Olmesartan on cartilage ECM degradation. Interleukin-29 (IL-29) is a novel inflammatory mediator involved in the inflammation and degradation of cartilage in OA, and human T/C-28a2 cells treated with it were the inflammatory model in vitro. We found that the degradation of type 2 collagens and aggrecans was induced by IL-29, accompanied by the upregulation of MMPs and ADAMTSs, but the presence of Olmesartan significantly ameliorated these increases. In addition, Olmesartan abolished IL-29- induced oxidative stress and elevated the expression level of TNF receptor-associated factor 6 (TRAF-6). Mechanistically, we showed that Olmesartan suppressed IL-29- caused inhibitor kappa B α (IκBα) expression and nuclear translocation of nuclear factor kappa-B (NF-κB) p65, indicating it suppressed the activation of the NF-κB pathway. Collectively, our data reveal that Olmesartan exerted a protective function on IL-29- induced type 2 collagen degradation in human chondrocytes.
- Published
- 2022
6. Long non-coding RNA H19 aggravates keloid progression by upregulating SMAD family member 5 expression via miR-196b-5p
- Author
-
Li, Zhichao, Gong, Cheng, and Wei, Huiming
- Subjects
Smad5 Protein ,Cytoplasm ,Cell Survival ,Primary Cell Culture ,Down-Regulation ,lncrna h19 ,Bioengineering ,Applied Microbiology and Biotechnology ,Cell Movement ,Humans ,smad5 ,skin and connective tissue diseases ,Cells, Cultured ,Cell Proliferation ,Cell Nucleus ,General Medicine ,Fibroblasts ,mir-196b-5p ,female genital diseases and pregnancy complications ,keloid ,MicroRNAs ,embryonic structures ,Disease Progression ,RNA, Long Noncoding ,TP248.13-248.65 ,Research Article ,Research Paper ,Biotechnology - Abstract
Accumulating evidence suggests that long non-coding RNAs (lncRNAs) participate in the formation and development of keloids, a benign tumor. In addition, lncRNA H19 has been shown to act on the biological processes of keloids. This study aimed to identify other important mechanisms of the effect of lncRNA H19 on keloid formation. The H19, miR-196b-5p, and SMAD family member 5 (SMAD5) expression levels were detected using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and Western blotting. Subcellular localization of lncRNA H19 was detected using a nuclear–cytoplasmic separation assay. Cell viability and proliferation were measured using counting kit-8 and colony formation assays. Bax and Bcl-2 levels were examined using Western blot analysis. The interaction between H19 and miR-196b-5p or SMAD5 was verified using a dual-luciferase reporter assay. H19 and SMAD5 expression was upregulated in keloid tissue and fibroblasts, whereas miR-196b-5p expression was downregulated. Knockdown of H19, overexpression of miR-196b-5p, or knockdown of SMAD5 inhibited the viability and proliferation of keloid fibroblasts and promoted apoptosis. Overexpression of H19 or SMAD5 and knockdown of miR-196b-5p promoted viability and proliferation and inhibited apoptosis. miR-196b-5p was identified as a H19 sponge, and SMAD5 was identified as a miR-196b-5p target. The combination of lncRNA H19 and miR-196b-5p regulates SMAD5 expression and promotes keloid formation, thus providing a new direction for keloid treatment.
- Published
- 2022
7. Protective effect of dexmedetomidine in cecal ligation perforation-induced acute lung injury through HMGB1/RAGE pathway regulation and pyroptosis activation
- Author
-
Huaqin Sun, Mingsun Fang, Hongyi Hu, Xiaoping Xu, Zhehao Liang, and Tao Tao
- Subjects
Male ,animal diseases ,Acute Lung Injury ,Receptor for Advanced Glycation End Products ,Perforation (oil well) ,Bioengineering ,Lung injury ,Pharmacology ,Protective Agents ,HMGB1 ,Applied Microbiology and Biotechnology ,NF-κB ,Cell Line ,RAGE (receptor) ,Intensive care ,polycyclic compounds ,Pyroptosis ,Animals ,Medicine ,RNA, Messenger ,HMGB1 Protein ,Cecum ,Ligation ,Lung ,Cell Nucleus ,biology ,business.industry ,Lentivirus ,RAGE ,General Medicine ,respiratory system ,respiratory tract diseases ,Mice, Inbred C57BL ,Disease Models, Animal ,Protein Transport ,Myeloperoxidase ,biology.protein ,Cytokines ,Tumor necrosis factor alpha ,Inflammation Mediators ,business ,TP248.13-248.65 ,hormones, hormone substitutes, and hormone antagonists ,Dexmedetomidine ,Research Article ,Research Paper ,Signal Transduction ,Biotechnology - Abstract
Dexmedetomidine (DEX) has been reported to attenuate cecal ligation perforation (CLP)-stimulated acute lung injury (ALI) by downregulating HMGB1 and RAGE. This study aimed to further investigate the specific mechanisms of RAGE and its potential-related mechanisms of DEX on ALI models in vitro and in vivo. The in vitro and in vivo ALI models were established by lipopolysaccharide treatment in MLE-12 cells and CLP in mice, respectively. The effect of DEX on pathological alteration was investigated by HE staining. Thereafter, the myeloperoxidase (MPO) activity and inflammatory cytokine levels were respectively detected to assess the lung injury of mice using commercial kits. The expression levels of HMGB1, RAGE, NF-κB, and pyroptosis-related molecules were detected by RT-qPCR and Western blot. HE staining showed that lung injury, increased inflammatory cell infiltration, and lung permeability was found in the ALI mice, and DEX treatment significantly attenuated lung tissue damage induced by CLP. The MPO activity and inflammatory cytokines (TNF-α, IL-1β, and NLRP3) levels were also significantly reduced after DEX treatment compared with those in the ALI mice. Moreover, DEX activated the HMGB1/RAGE/NF-κB pathway and upregulated the pyroptosis-related proteins. However, the protective DEX effect was impaired by RAGE overexpression in ALI mice and MLE-12 cells. Additionally, DEX treatment significantly suppressed HMGB1 translocation from the nucleus region to the cytoplasm, and this effect was reversed by RAGE overexpression. These findings suggested that DEX may be a useful ALI treatment, and the protective effects on ALI mice may be through the inhibition of HMGB1/RAGE/NF-κB pathway and cell pyroptosis., Graphical abstractDexmedetomidine (DEX) has protective effects on acute lung injury (ALI) in vitro and in vivo. The possible mechanisms may be closely associated with inflammatory response, caspase-1-mediated cell pyroptosis, and high-mobility group protein 1 (HMGB1)/receptor for advanced glycation end products (RAGE)/nuclear factor-κB pathway. Moreover, DEX could promote the HMGB1 translocation from the cytoplasm to the nucleus in lipopolysaccharide-activated MLE-12 cells, whereas the action of RAGE overexpression was opposite.
- Published
- 2021
8. Curcumin protection against ultraviolet-induced photo-damage in Hacat cells by regulating nuclear factor erythroid 2-related factor 2
- Author
-
Runxiang Li, Huilan Zhu, Miaojian Wan, Huiyan Deng, Huaping Li, Quan Chen, and Bihua Liang
- Subjects
Keratinocytes ,skin ,Cell Survival ,NF-E2-Related Factor 2 ,Ultraviolet Rays ,Apoptosis ,Bioengineering ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Antioxidants ,Superoxide dismutase ,chemistry.chemical_compound ,ultraviolet ,medicine ,HaCaT Cells ,Humans ,oxidative stress ,curcumin ,Cell Proliferation ,Cell Nucleus ,biology ,integumentary system ,Cell growth ,Chemistry ,photo-damage ,General Medicine ,respiratory system ,Molecular biology ,Heme oxygenase ,HaCaT ,nuclear factor erythroid 2-related factor 2 ,Cytoprotection ,Catalase ,biology.protein ,Curcumin ,Oxidative stress ,TP248.13-248.65 ,Research Article ,Research Paper ,Biotechnology - Abstract
Curcumin suppressed ultraviolet (UV) induced skin carcinogenesis and activated the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. However, whether curcumin protects skin injury caused by UV is still unknown. A vitro model was established and curcumin effects on Hacat cells were detected. Nrf2 was knocked down in Hacat cells to verify the Nrf2 role in the protective effect of curcumin. Results indicated that ultraviolet A (UVA) (or ultraviolet B (UVB)) irradiation would lead to decreased cell proliferation, increased cell apoptosis, decreased catalase, heme oxygenase 1, and superoxide dismutase expression, and increased levels of protein carbonylation and malondialdehyde (p
- Published
- 2021
9. Nucleo-cytoplasmic RNA distribution responsible for maintaining neuroinflammatory microenvironment
- Author
-
Jianhua Peng, Xiancheng Qiu, Yuyan Liao, Chenghao Kuang, Long Gu, Lifang Zhang, Shigang Yin, Xi Kong, Yong Jiang, Ghosh Dipritu, Qianke Tao, Zheng Bao, and Yijing He
- Subjects
Cytoplasm ,Hemorrhage ,Biology ,RNA Transport ,Cell Line ,Mice ,medicine ,Animals ,Molecular Biology ,Gene ,Neuroinflammation ,Cell Nucleus ,Regulation of gene expression ,Microglia ,Gene Expression Profiling ,Computational Biology ,RNA ,Cell Biology ,Subcellular localization ,Antisense RNA ,Cell biology ,Disease Models, Animal ,Gene Ontology ,medicine.anatomical_structure ,Cellular Microenvironment ,Neuroinflammatory Diseases ,RNA, Long Noncoding ,Disease Susceptibility ,Research Paper - Abstract
Subcellular localization of transcripts is highly associated with regulation of gene expression, synthesis of protein, and also the development of the human brain cortex. Although many mechanisms are prevalent in the occurrence of neuroinflammation, the mechanisms based on differences in subcellular localization of transcripts have not been explored. To characterize the dynamic profile of nuclear and cytoplasmic transcripts during the progress of haemorrhage-induced neuroinflammation, we isolated nucleo-cytoplasmic RNA fractions of oxyhaemoglobin (oxy-Hb) treated microglia cells and sequenced both fractions. We discovered that cytoplasmic retained genes were the major forces to maintain the neuroinflammatory microenvironment with 10 hub genes and 40 conserved genes were identified. Moreover, antisense RNA Gm44096 and lincRNA Gm47270, which co-expressed with a crowd of inflammatory genes in the cytoplasm, were discovered as regulatory strategies for sustaining the neuroinflammatory microenvironment. Thus, our study provides a new perspective on understanding haemorrhage-induced neuroinflammation and also reveals a mechanism of lncRNA responsible for maintaining the neuroinflammatory microenvironment.
- Published
- 2021
10. Challenges of Using Expansion Microscopy for Super‐resolved Imaging of Cellular Organelles
- Author
-
Christoffer Lagerholm, Christian Eggeling, Silvia Galiani, Dominic Waithe, Wolfgang Schliebs, Katharina Reglinski, Ralf Erdmann, and Maximilian Büttner
- Subjects
cell organelles ,Matrix (biology) ,010402 general chemistry ,01 natural sciences ,Biochemistry ,law.invention ,Very Important Paper ,STED microscopy ,Confocal microscopy ,law ,Microscopy ,Organelle ,Peroxisomes ,Humans ,bioorganic chemistry ,Molecular Biology ,Polyacrylamide gel electrophoresis ,Cellular compartment ,Cell Nucleus ,Microscopy, Confocal ,Full Paper ,010405 organic chemistry ,Chemistry ,Cell Membrane ,Organic Chemistry ,Full Papers ,Peroxisome ,Mitochondria ,Molecular Imaging ,0104 chemical sciences ,HEK293 Cells ,Microscopy, Fluorescence ,peptides ,Biophysics ,Molecular Medicine ,expansion microscopy - Abstract
Expansion microscopy (ExM) has been successfully used to improve the spatial resolution when imaging tissues by optical microscopy. In ExM, proteins of a fixed sample are crosslinked to a swellable acrylamide gel, which expands when incubated in water. Therefore, ExM allows enlarged subcellular structures to be resolved that would otherwise be hidden to standard confocal microscopy. Herein, we aim to validate ExM for the study of peroxisomes, mitochondria, nuclei and the plasma membrane. Upon comparison of the expansion factors of these cellular compartments in HEK293 cells within the same gel, we found significant differences, of a factor of above 2, in expansion factors. For peroxisomes, the expansion factor differed even between peroxisomal membrane and matrix marker; this underlines the need for a thorough validation of expansion factors of this powerful technique. We further give an overview of possible quantification methods for the determination of expansion factors of intracellular organelles, and we highlight some potentials and challenges., In Expansion Microscopy (ExM) subcellular structures are imaged in isotropically expanded fixed samples, consequently allowing to resolve enlarged subcellular structures, otherwise hidden to standard microscopy. Upon comparison of the expansion factors of different cellular compartments in cells within the same gel, we found significant differences in expansion factors of a factor of above 2.
- Published
- 2020
11. The photoreactivation of 6 - 4 photoproducts in chloroplast and nuclear DNA depends on the amount of the Arabidopsis UV repair defective 3 protein.
- Author
-
Zgłobicki P, Hermanowicz P, Kłodawska K, Bażant A, Łabuz J, Grzyb J, Dutka M, Kowalska E, Jawor J, Leja K, and Banaś AK
- Subjects
- Ultraviolet Rays, DNA, Plant metabolism, DNA, Plant genetics, Pyrimidine Dimers metabolism, Pyrimidine Dimers genetics, DNA, Chloroplast genetics, DNA, Chloroplast metabolism, Chloroplasts metabolism, DNA Damage, Arabidopsis genetics, Arabidopsis radiation effects, Arabidopsis metabolism, Arabidopsis Proteins metabolism, Arabidopsis Proteins genetics, Cell Nucleus metabolism, Cell Nucleus radiation effects, Deoxyribodipyrimidine Photo-Lyase metabolism, Deoxyribodipyrimidine Photo-Lyase genetics, Plants, Genetically Modified, DNA Repair
- Abstract
Background: 6 - 4 photoproducts are the second most common UV-induced DNA lesions after cyclobutane pyrimidine dimers. In plants, they are mainly repaired by photolyases in a process called photoreactivation. While pyrimidine dimers can be deleterious, leading to mutagenesis or even cell death, 6 - 4 photoproducts can activate specific signaling pathways. Therefore, their removal is particularly important, especially for plants exposed to high UV intensities due to their sessile nature. Although photoreactivation in nuclear DNA is well-known, its role in plant organelles remains unclear. In this paper we analyzed the activity and localization of GFP-tagged AtUVR3, the 6 - 4 photoproduct specific photolyase., Results: Using transgenic Arabidopsis with different expression levels of AtUVR3, we confirmed a positive trend between these levels and the rate of 6 - 4 photoproduct removal under blue light. Measurements of 6 - 4 photoproduct levels in chloroplast and nuclear DNA of wild type, photolyase mutants, and transgenic plants overexpressing AtUVR3 showed that the photoreactivation is the main repair pathway responsible for the removal of these lesions in both organelles. The GFP-tagged AtUVR3 was predominantly located in nuclei with a small fraction present in chloroplasts and mitochondria of transgenic Arabidopsis thaliana and Nicotiana tabacum lines. In chloroplasts, this photolyase co-localized with the nucleoid marked by plastid envelope DNA binding protein., Conclusions: Photolyases are mainly localized in plant nuclei, with only a small fraction present in chloroplasts and mitochondria. Despite this unbalanced distribution, photoreactivation is the primary mechanism responsible for the removal of 6 - 4 photoproducts from nuclear and chloroplast DNA in adult leaves. The amount of the AtUVR3 photolyase is the limiting factor influencing the photoreactivation rate of 6 - 4 photoproducts. The efficient photoreactivation of 6 - 4 photoproducts in 35S: AtUVR3-GFP Arabidopsis and Nicotiana tabacum is a promising starting point to evaluate whether transgenic crops overproducing this photolyase are more tolerant to high UV irradiation and how they respond to other abiotic and biotic stresses under field conditions., (© 2024. The Author(s).)
- Published
- 2024
- Full Text
- View/download PDF
12. FAK, vinculin, and talin control mechanosensitive YAP nuclear localization.
- Author
-
Holland EN, Fernández-Yagüe MA, Zhou DW, O'Neill EB, Woodfolk AU, Mora-Boza A, Fu J, Schlaepfer DD, and García AJ
- Subjects
- Humans, Adaptor Proteins, Signal Transducing metabolism, Transcription Factors metabolism, Mesenchymal Stem Cells metabolism, Mesenchymal Stem Cells cytology, Animals, Focal Adhesions metabolism, Mice, Fibroblasts metabolism, Focal Adhesion Kinase 1 metabolism, Focal Adhesion Protein-Tyrosine Kinases metabolism, Protein Binding, Talin metabolism, Vinculin metabolism, Cell Nucleus metabolism, YAP-Signaling Proteins metabolism, Mechanotransduction, Cellular
- Abstract
Focal adhesions (FAs) are nanoscale complexes containing clustered integrin receptors and intracellular structural and signaling proteins that function as principal sites of mechanotransduction in part via promoting the nuclear translocation and activation of the transcriptional coactivator yes-associated protein (YAP). Knockdown of FA proteins such as focal adhesion kinase (FAK), talin, and vinculin can prevent YAP nuclear localization. However, the mechanism(s) of action remain poorly understood. Herein, we investigated the role of different functional domains in vinculin, talin, and FAK in regulating YAP nuclear localization. Using genetic or pharmacological inhibition of fibroblasts and human mesenchymal stem cells (hMSCs) adhering to deformable substrates, we find that disruption of vinculin-talin binding versus talin-FAK binding reduces YAP nuclear localization and transcriptional activity via different mechanisms. Disruption of vinculin-talin binding or knockdown of talin-1 reduces nuclear size, traction forces, and YAP nuclear localization. In contrast, disruption of the talin binding site on FAK or elimination of FAK catalytic activity did not alter nuclear size yet still prevented YAP nuclear localization and activity. These data support both nuclear tension-dependent and independent models for matrix stiffness-regulated YAP nuclear localization. Our results highlight the importance of vinculin-talin-FAK interactions at FAs of adherent cells, controlling YAP nuclear localization and activity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)
- Published
- 2024
- Full Text
- View/download PDF
13. Re-assessment of the subcellular localization of Bazooka/Par-3 in Drosophila: no evidence for localization to the nucleus and the neuromuscular junction.
- Author
-
Kim S, Shahab J, Vogelsang E, and Wodarz A
- Subjects
- Animals, Protein Transport, Cell Nucleus metabolism, Drosophila Proteins metabolism, Drosophila Proteins genetics, Neuromuscular Junction metabolism, Drosophila melanogaster genetics, Drosophila melanogaster metabolism
- Abstract
Bazooka/Par-3 (Baz) is an evolutionarily conserved scaffold protein that functions as a master regulator for the establishment and maintenance of cell polarity in many different cell types. In the vast majority of published research papers Baz has been reported to localize at the cell cortex and at intercellular junctions. However, there have also been several reports showing localization and function of Baz at additional subcellular sites, in particular the nuclear envelope and the neuromuscular junction. In this study we have re-assessed the localization of Baz to these subcellular sites in a systematic manner. We used antibodies raised in different host animals against different epitopes of Baz for confocal imaging of Drosophila tissues. We tested the specificity of these antisera by mosaic analysis with null mutant baz alleles and tissue-specific RNAi against baz. In addition, we used a GFP-tagged gene trap line for Baz and a bacterial artificial chromosome (BAC) expressing GFP-tagged Baz under control of its endogenous promoter in a baz mutant background to compare the subcellular localization of the GFP-Baz fusion proteins to the staining with anti-Baz antisera. Together, these experiments did not provide evidence for specific localization of Baz to the nucleus or the neuromuscular junction., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)
- Published
- 2024
- Full Text
- View/download PDF
14. BoNuS: Boundary Mining for Nuclei Segmentation With Partial Point Labels.
- Author
-
Lin Y, Wang Z, Zhang D, Cheng KT, and Chen H
- Subjects
- Humans, Image Processing, Computer-Assisted methods, Databases, Factual, Image Interpretation, Computer-Assisted methods, Cell Nucleus, Algorithms
- Abstract
Nuclei segmentation is a fundamental prerequisite in the digital pathology workflow. The development of automated methods for nuclei segmentation enables quantitative analysis of the wide existence and large variances in nuclei morphometry in histopathology images. However, manual annotation of tens of thousands of nuclei is tedious and time-consuming, which requires significant amount of human effort and domain-specific expertise. To alleviate this problem, in this paper, we propose a weakly-supervised nuclei segmentation method that only requires partial point labels of nuclei. Specifically, we propose a novel boundary mining framework for nuclei segmentation, named BoNuS, which simultaneously learns nuclei interior and boundary information from the point labels. To achieve this goal, we propose a novel boundary mining loss, which guides the model to learn the boundary information by exploring the pairwise pixel affinity in a multiple-instance learning manner. Then, we consider a more challenging problem, i.e., partial point label, where we propose a nuclei detection module with curriculum learning to detect the missing nuclei with prior morphological knowledge. The proposed method is validated on three public datasets, MoNuSeg, CPM, and CoNIC datasets. Experimental results demonstrate the superior performance of our method to the state-of-the-art weakly-supervised nuclei segmentation methods. Code: https://github.com/hust-linyi/bonus.
- Published
- 2024
- Full Text
- View/download PDF
15. The plant nuclear lamina proteins NMCP1 and NMCP2 form a filamentous network with lateral filament associations
- Author
-
Kaien Fujino, Kiyoshi Masuda, and Riku Hikida
- Subjects
0106 biological sciences ,NMCP2 ,NMCP1 ,Physiology ,Plant Science ,macromolecular substances ,Immunofluorescence ,01 natural sciences ,Protein filament ,03 medical and health sciences ,Nuclear Matrix-Associated Proteins ,stimulated emission depletion microscopy ,super-resolution microscopy ,parasitic diseases ,medicine ,Coiled-coil protein ,natural sciences ,Intermediate filament ,030304 developmental biology ,Plant Proteins ,Cell Nucleus ,0303 health sciences ,filamentous network ,medicine.diagnostic_test ,Chemistry ,Super-resolution microscopy ,AcademicSubjects/SCI01210 ,technology, industry, and agriculture ,food and beverages ,Cell Biology ,self-assembly ,Nuclear matrix ,Research Papers ,Lamins ,Chromatin ,Biophysics ,Nuclear lamina ,Lamin ,nuclear lamina ,010606 plant biology & botany - Abstract
Super-resolution microscopy revealed that the plant nuclear lamina proteins NMCP1 and NMCP2 organize into filamentous networks with filament bundles., Plant genomes lack genes encoding intermediate filament proteins, including lamins; however, functional lamin analogues are presumed to exist in plants. Plant-specific coiled-coil proteins, that is, nuclear matrix constituent proteins (NMCPs), are the most likely candidates as the structural elements of the nuclear lamina because they exhibit a lamin-like domain arrangement. They are exclusively localized at the nuclear periphery and have functions that are analogous to those of lamins. However, their assembly into filamentous polymers has not yet been confirmed. In this study, we examined the higher-order structure of NMCP1 and NMCP2 in Apium graveolens cells by using stimulated emission depletion microscopy combined with immunofluorescence cell labelling. Our analyses revealed that NMCP1 and NMCP2 form intricate filamentous networks, which include thick segments consisting of filament bundles, forming a dense filamentous layer extending across the nuclear periphery. Furthermore, the outermost chromatin distribution was found to be in the nucleoplasm-facing region of the nuclear lamina. Recombinant Daucus carota NMCP1 with a His-tag produced in Escherichia coli refolded into dimers and self-assembled into filaments and filament bundles. These results suggest that NMCP1 and NMCP2 organize into the nuclear lamina by forming a filamentous network with filament bundles that localize at the nuclear periphery.
- Published
- 2021
16. Nuclear expression of VDR and AHR is mutually exclusive in glandular cells in endometriosis
- Author
-
Livio Casarini, Viktoria von Schönfeldt, Francesco De Pascali, Manuela Simoni, Sven Mahner, Christina Kuhn, and Udo Jeschke
- Subjects
0301 basic medicine ,Histology ,AHR ,Response element ,Endometriosis ,Biology ,Endometrium ,Calcitriol receptor ,03 medical and health sciences ,Immunohistochemistry ,Nuclear receptors ,VDR ,0302 clinical medicine ,Basic Helix-Loop-Helix Transcription Factors ,medicine ,Humans ,Receptor ,Molecular Biology ,Cell Nucleus ,Original Paper ,030219 obstetrics & reproductive medicine ,Epithelial Cells ,Cell Biology ,medicine.disease ,Aryl hydrocarbon receptor ,Medical Laboratory Technology ,030104 developmental biology ,medicine.anatomical_structure ,Receptors, Aryl Hydrocarbon ,Nuclear receptor ,Cancer research ,biology.protein ,Receptors, Calcitriol ,Female - Abstract
The vitamin D receptor (VDR) and aryl hydrocarbon receptor (AHR) are two nuclear receptors that exert their effects by binding with ligands and forming a molecular complex. These complexes translocate to the nucleus and activate the expression of a series of genes which have a response element to VDR or AHR. Both receptors have been identified in the pathogenesis of endometriosis, a common disease characterized by the formation of endometrium-like tissue in ectopic zones. Despite numerous therapies, there is no definitive cure for endometriosis at the pharmacological level. Our study aims to describe the location and the expression of VDR and AHR at the protein level. For this purpose, an evaluation was performed using tissue from the three normal phases of the endometrium (proliferative, early, and late secretory) and in endometriosis by immunohistochemistry, using anti-VDR and anti-AHR antibodies. We demonstrate that in the nuclei of glandular cells in endometriosis, the expression of VDR and AHR is mutually exclusive—when the expression of one receptor is high, the other one is low—suggesting a possible target in the treatment of endometriosis. We also identify a significant change in the expression of glandular cytoplasmic AHR between the proliferative and late secretory endometrium. Supplementary Information The online version contains supplementary material available at 10.1007/s00418-021-02005-9.
- Published
- 2021
17. PTEN nuclear translocation enhances neuronal injury after hypoxia-ischemia via modulation of the nuclear factor-κB signaling pathway
- Author
-
Chunyan Zhang, Jing Zhao, Li Hou, Lin Jiang, Linlin Yin, and Ling He
- Subjects
Aging ,MAP Kinase Signaling System ,Phosphatase ,Brain damage ,Rats, Sprague-Dawley ,Ischemia ,medicine ,PTEN ,Tensin ,Animals ,Phosphorylation ,Cells, Cultured ,Cell Nucleus ,Neurons ,biology ,ERK1/2 ,Kinase ,Chemistry ,oxygen and glucose deprivation ,NF-kappa B ,PTEN Phosphohydrolase ,Cell Biology ,brain damage ,Cell Hypoxia ,Cell biology ,Oxygen ,hypoxia-ischemia ,Protein Transport ,phosphatase and tensin homolog deletion from chromosome 10 ,Glucose ,nervous system ,Apoptosis ,biology.protein ,medicine.symptom ,Signal transduction ,Research Paper ,Signal Transduction - Abstract
The occurrence of hypoxia-ischemia (HI) in the developing brain is closely associated with neuronal injury and even death. However, the underlying molecular mechanism is not fully understood. This study was designed to investigate phosphatase and tensin homolog (PTEN) nuclear translocation and its possible role in rat cortical neuronal damage following oxygen-glucose deprivation (OGD) in vitro. An in vitro OGD model was established using primary cortical neurons dissected from newborn Sprague-Dawley rats to mimic HI conditions. The PTENK13R mutant plasmid, which contains a lysine-to-arginine mutation at the lysine 13 residue, was constructed. The nuclei and cytoplasm of neurons were separated. Neuronal injury following OGD was evidenced by increased lactate dehydrogenase (LDH) release and apoptotic cell counts. In addition, PTEN expression was increased and the phosphorylation of extracellular signal-regulated kinase 1/2 (p-ERK1/2) and activation of nuclear factor kappa B (NF-κB) were decreased following OGD. PTENK13R transfection prevented PTEN nuclear translocation; attenuated the effect of OGD on nuclear p-ERK1/2 and NF-κB, apoptosis, and LDH release; and increased the expression of several anti-apoptotic proteins. We conclude that PTEN nuclear translocation plays an essential role in neuronal injury following OGD via modulation of the p-ERK1/2 and NF-κB pathways. Prevention of PTEN nuclear translocation might be a candidate strategy for preventing brain injury following HI.
- Published
- 2021
18. Racemization in cataractous lens from diabetic and aging individuals: analysis of Asp 58 residue in αA-crystallin
- Author
-
Jiao Qi, Wenwen He, Keke Zhang, Xiangjia Zhu, and Yi Lu
- Subjects
Male ,medicine.medical_specialty ,Aging ,genetic structures ,lens ,Cataract ,Lens protein ,Isomerism ,Crystallin ,Cortex (anatomy) ,Internal medicine ,Diabetes mellitus ,crystallin ,Lens, Crystalline ,medicine ,Humans ,aspartyl residue ,Racemization ,Aged ,Cell Nucleus ,Aspartic Acid ,Arc (protein) ,diabetes ,Chemistry ,Cell Biology ,medicine.disease ,Crystallins ,eye diseases ,medicine.anatomical_structure ,Endocrinology ,Solubility ,racemization ,Cataractous lens ,Female ,sense organs ,Nucleus ,Research Paper - Abstract
Cataract is the leading cause of visual impairment globally. Racemization of lens proteins may contribute to cataract formation in aging individuals. As a special type of age-related cataract (ARC), diabetic cataract (DC) is characterized by the early onset of cortical opacification and finally developed into a mixed type of cortical and nuclear opacification. We compared racemization of Asp 58 residue, a hotspot position in αA-crystallin, from the cortex and nucleus of diabetic and age-matched senile cataractous lenses, by identifying L-Asp/L-isoAsp/D-Asp/D-isoAsp by mass spectrometry. Compared to nondiabetic cataractous lenses, DC lenses showed a significantly increased cortex/nucleus ratio of D-Asp 58, which originated primarily from an increased percentage of D-Asp 58 in the lens cortex of DC. Moreover, patients diagnosed with diabetes for over 10 years showed a lower cortex/nucleus ratio of D-isoAsp 58 in the lens compared with those who had a shorter duration of diabetes, which originated mainly from an increased percentage of D-isoAsp 58 in the lens nucleus of DC with increasing time of hyperglycemia. Further analysis confirmed decreased protein solubility in diabetic cataractous lenses. The different racemization pattern in DC may be distinguished from ARC and influence its phenotype over the protracted duration of diabetes.
- Published
- 2021
19. Inhibition of CXCR2 plays a pivotal role in re-sensitizing ovarian cancer to cisplatin treatment
- Author
-
Nayara G. Tessarollo, Raquel Maria da Silva Graça Almeida, Luísa Pereira, Diandra Zipinotti dos Santos, Ian Victor Silva, Patrícia Mesquita, Isabella dos Santos Guimarães, Diana Pádua, Paulo Cilas Morais Lyra-Junior, Leticia Batista Azevedo Rangel, Taciane B. Henriques, Ana Luísa Amaral, and Bruno Cavadas
- Subjects
Aging ,Epithelial-Mesenchymal Transition ,Cell Survival ,MAP Kinase Signaling System ,Chemokine CXCL2 ,Chick Embryo ,Receptors, Interleukin-8B ,Phosphatidylinositol 3-Kinases ,Chemokine receptor ,Cell Line, Tumor ,medicine ,Animals ,Humans ,tumor microenvironment ,Neoplasm Invasiveness ,Interleukin 8 ,CXC chemokine receptors ,Cell Proliferation ,Cell Nucleus ,Ovarian Neoplasms ,Cisplatin ,Tumor microenvironment ,CXCR2 ,Neovascularization, Pathologic ,Chemistry ,TOR Serine-Threonine Kinases ,chemoresistance ,Cell Biology ,medicine.disease ,Survival Analysis ,Neoplasm Proteins ,CXCL2 ,Drug Resistance, Neoplasm ,Tumor progression ,high grade serous ovarian cancer ,Cancer research ,Female ,Ovarian cancer ,Proto-Oncogene Proteins c-akt ,Research Paper ,medicine.drug - Abstract
cDNA microarray data conducted by our group revealed overexpression of CXCL2 and CXCL8 in ovarian cancer (OC) microenvironment. Herein, we have proven that the chemokine receptor, CXCR2, is a pivotal molecule in re-sensitizing OC to cisplatin, and its inhibition decreases cell proliferation, viability, tumor size in cisplatin-resistant cells, as well as reversed the overexpression of mesenchymal epithelium transition markers. Altogether, our study indicates a central effect of CXCR2 in preventing tumor progression, due to acquisition of cisplatin chemoresistant phenotype by tumor cells, and patients’ high lethality rate. We found that the overexpression of CXCR2 by OC cells is persistent and anomalously confined to the cellular nuclei, thus pointing to an urge in developing highly lipophilic molecules that promptly permeate cells, bind to and inhibit nuclear CXCR2 to fight OC, instead of relying on the high-cost genetic engineered cells.
- Published
- 2021
20. SARS-CoV-2 main protease suppresses type I interferon production by preventing nuclear translocation of phosphorylated IRF3
- Author
-
Sin-Yee Fung, Dong-Yan Jin, Huayue Lin, Man Lung Yeung, and Kam-Leung Siu
- Subjects
medicine.medical_treatment ,viruses ,NSP5 ,Applied Microbiology and Biotechnology ,03 medical and health sciences ,TANK-binding kinase 1 ,Interferon ,Chlorocebus aethiops ,medicine ,Animals ,Humans ,Phosphorylation ,skin and connective tissue diseases ,Molecular Biology ,Vero Cells ,Ecology, Evolution, Behavior and Systematics ,Coronavirus 3C Proteases ,030304 developmental biology ,Cell Nucleus ,0303 health sciences ,Protease ,Chemistry ,SARS-CoV-2 ,fungi ,virus diseases ,COVID-19 ,SARS-CoV ,Cell Biology ,biochemical phenomena, metabolism, and nutrition ,IRF3 ,Type I interferon production ,Cell biology ,Protein Transport ,type I interferons ,Interferon Type I ,Vero cell ,Interferon Regulatory Factor-3 ,3C-like protease ,Interferon type I ,Developmental Biology ,medicine.drug ,Research Paper - Abstract
Suppression of type I interferon (IFN) response is one pathological outcome of the infection of highly pathogenic human coronaviruses. To effect this, severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2 encode multiple IFN antagonists. In this study, we reported on the IFN antagonism of SARS-CoV-2 main protease NSP5. NSP5 proteins of both SARS-CoV and SARS-CoV-2 counteracted Sendai virus-induced IFN production. NSP5 variants G15S and K90R commonly seen in circulating strains of SARS-CoV-2 retained the IFN-antagonizing property. The suppressive effect of NSP5 on IFN-β gene transcription induced by RIG-I, MAVS, TBK1 and IKKϵ suggested that NSP5 likely acts at a step downstream of IRF3 phosphorylation in the cytoplasm. NSP5 did not influence steady-state expression or phosphorylation of IRF3, suggesting that IRF3, regardless of its phosphorylation state, might not be the substrate of NSP5 protease. However, nuclear translocation of phosphorylated IRF3 was severely compromised in NSP5-expressing cells. Taken together, our work revealed a new mechanism by which NSP5 proteins encoded by SARS-CoV and SARS-CoV-2 antagonize IFN production by retaining phosphorylated IRF3 in the cytoplasm. Our findings have implications in rational design and development of antiviral agents against SARS-CoV-2.
- Published
- 2021
21. The functional impact of nuclear reorganization in cellular senescence
- Author
-
Azucena Rocha, Audrey Dalgarno, and Nicola Neretti
- Subjects
Cell Nucleus ,AcademicSubjects/SCI01140 ,Review Paper ,0303 health sciences ,nuclear organization ,aging ,epigenome ,Cellular senescence ,Functional impact ,Genomics ,General Medicine ,Biology ,Biochemistry ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,Genetics ,cellular senescence ,chromatin ,Molecular Biology ,030217 neurology & neurosurgery ,030304 developmental biology - Abstract
Cellular senescence is the irreversible cell cycle arrest in response to DNA damage. Because senescent cells accumulate with age and contribute to chronic inflammation, they are promising therapeutic targets for healthspan extension. The senescent phenotype can vary depending on cell type and on the specific insults that induce senescence. This variability is also reflected in the extensive remodeling of the genome organization within the nucleus of senescent cells. Here, we give an overview of the nuclear changes that occur in different forms of senescence, including changes to chromatin state and composition and to the three-dimensional organization of the genome, as well as alterations to the nuclear envelope and to the accessibility of repetitive genomic regions. Many of these changes are shared across all forms of senescence, implicating nuclear organization as a fundamental driver of the senescent state and of how senescent cells interact with the surrounding tissue.
- Published
- 2021
22. Coxsackievirus B3 targets TFEB to disrupt lysosomal function
- Author
-
Chen Seng Ng, Amirhossein Bahreyni, Yuanchao Xue, Honglin Luo, Yasir Mohamud, Hui Tang, and Huitao Liu
- Subjects
0301 basic medicine ,autophagy ,Viral Myocarditis ,3C proteinase ,viruses ,Biology ,medicine.disease_cause ,03 medical and health sciences ,Lysosome ,medicine ,Humans ,Coxsackievirus B3 ,Molecular Biology ,Cell Nucleus ,TFEB ,030102 biochemistry & molecular biology ,enterovirus ,Basic Helix-Loop-Helix Leucine Zipper Transcription Factors ,Autophagy ,virus diseases ,Cell Biology ,Virology ,Enterovirus B, Human ,3. Good health ,Protein Transport ,030104 developmental biology ,medicine.anatomical_structure ,Coxsackievirus b3 ,lysosome ,cardiovascular system ,Enterovirus ,Lysosomes ,Function (biology) ,Research Article ,Research Paper - Abstract
Coxsackievirus B3 (CVB3) is a prevalent etiological agent for viral myocarditis and neurological disorders, particularly in infants and young children. Virus-encoded proteinases have emerged as cytopathic factors that contribute to disease pathogenesis in part through targeting the cellular recycling machinery of autophagy. Although it is appreciated that CVB3 can usurp cellular macroautophagy/autophagy for pro-viral functions, the precise mechanisms by which viral proteinases disrupt autophagy remain incompletely understood. Here we identified TFEB (transcription factor EB), a master regulator of autophagy and lysosome biogenesis, as a novel target of CVB3 proteinase 3 C. Time-course infections uncovered a significant loss of full-length TFEB and the emergence of a lower-molecular mass (~63 kDa) fragment. Cellular and in vitro cleavage assays revealed the involvement of viral proteinase 3 C in the proteolytic processing of TFEB, while site-directed mutagenesis identified the site of cleavage after glutamine 60. Assessment of TFEB transcriptional activity using a reporter construct discovered a loss of function of the cleavage fragment despite nuclear localization and retaining of the ability of DNA and protein binding. Furthermore, we showed that CVB3 infection was also able to trigger cleavage-independent nuclear translocation of TFEB that relied on the serine-threonine phosphatase PPP3/calcineurin. Finally, we demonstrated that both TFEB and TFEB [Δ60] serve roles in viral egress albeit through differing mechanisms. Collectively, this study reveals that CVB3 targets TFEB for proteolytic processing to disrupt host lysosomal function and enhance viral infection. Abbreviations:ACTB: actin beta; CLEAR: coordinated lysosomal enhancement and regulation; CVB3: coxsackievirus B3; DAPI: 4′,6-diamidino-2-phenylindole; GFP: green fluorescent protein; LAMP1: lysosomal associated membrane protein 1; LTR: LysoTracker Red; PPP3/calcineurin: protein phosphatase 3; PPP3CA: protein phosphatase 3 catalytic subunit A; p-TFEB: phospho-Ser211 TFEB; si-CON: scramble control siRNA; TFEB: transcription factor EB; TFEB [Δ60]: TFEB cleavage fragment that lacks the first 60 amino acids; VP1: viral capsid protein 1
- Published
- 2021
23. N6-methyladenosine-induced circ1662 promotes metastasis of colorectal cancer by accelerating YAP1 nuclear localization
- Author
-
Chen Chen, Weiwei Wang, Yuying Guo, Weitang Yuan, Zhenyu Ji, Luyang Zhao, Zhenqiang Sun, Guixian Wang, Xiuxiu Yang, Yaxin Guo, Qin Dang, Bo Shao, Jinbo Liu, Qiaozhen Kang, Shuaixi Yang, and Quanbo Zhou
- Subjects
0301 basic medicine ,Male ,Adenosine ,Colorectal cancer ,Medicine (miscellaneous) ,Metastasis ,Mice ,0302 clinical medicine ,Cell Movement ,Pharmacology, Toxicology and Pharmaceutics (miscellaneous) ,YAP1 ,Mice, Inbred BALB C ,Prognosis ,Gene Expression Regulation, Neoplastic ,030220 oncology & carcinogenesis ,Immunohistochemistry ,Female ,Colorectal Neoplasms ,Research Paper ,Signal Transduction ,circ1662 ,Mice, Nude ,colorectal cancer ,Biology ,03 medical and health sciences ,Cell Line, Tumor ,N6-methyladenosine (m6A) ,medicine ,Biomarkers, Tumor ,Animals ,Humans ,Smad3 Protein ,Adaptor Proteins, Signal Transducing ,Aged ,Cell Nucleus ,Messenger RNA ,RNA ,Cancer ,YAP-Signaling Proteins ,Methyltransferases ,RNA, Circular ,medicine.disease ,HCT116 Cells ,digestive system diseases ,030104 developmental biology ,HEK293 Cells ,Cancer research ,METTL3 ,Nuclear localization sequence ,Transcription Factors - Abstract
Tumor metastasis is the leading cause of death in patients with colorectal cancer (CRC). Circular RNAs (circRNAs) have been shown to be involved in cancer progression. However, the regulatory mechanisms of circRNAs involved in CRC tumor metastasis are currently unknown. Methods: High-throughput sequencing was performed on 6 pairs of CRC and adjacent normal tissues to identify the expression profiles of mRNA and circRNA. circ1662 was assessed by RNA-ISH and IHC of a tissue chip. The function of circ1662 in CRC was evaluated by knocking down or overexpressing circ1662. MeRIP-qPCR, RIP-qPCR, and RNA pull-down were performed to determine the relationship between METTL3, circ1662, and YAP1. Results: A novel circRNA, circ1662, exhibited significantly higher expression in CRC tissues than paired normal tissues. High circ1662 expression was correlated with poor prognosis and tumor depth in patients with CRC. Functionally, circ1662 promoted CRC cell invasion and migration by controlling EMT in vitro and in vivo. Mechanistically, circ1662 directly bound to YAP1 and accelerated its nuclear accumulation to regulate the SMAD3 pathway. Additionally, circ1662 enhanced CRC invasion and migration depending on YAP1 and SMAD3. Interestingly, METTL3 induced circ1662 expression by binding its flanking sequences and installing m6A modifications. Clinically, circ1662 expression strongly correlated with METTL3 and YAP1 protein expression. Moreover, YAP1 expression was negatively correlated with SMAD3 expression. Conclusions: METTL3-induced circ1662 promoted CRC cell invasion and migration by accelerating YAP1 nuclear transport. This result implies that circ1662 is a new prognostic and therapeutic marker for CRC metastasis.
- Published
- 2021
24. Ultrastructural observations on the oncomiracidium epidermis and adult tegument of Discocotyle sagittata, a monogenean gill parasite of salmonids
- Author
-
Jo Cable, Richard C. Tinsley, and Mohamed Mohamed El-Naggar
- Subjects
Gills ,Cytoplasm ,030231 tropical medicine ,Trematode Infections ,Aquaculture ,030308 mycology & parasitology ,Fish Diseases ,03 medical and health sciences ,0302 clinical medicine ,Animals ,Fish Parasitology - Original Paper ,Parasite hosting ,Cell Nucleus ,0303 health sciences ,Syncytium ,General Veterinary ,biology ,Embryogenesis ,Salmonids ,General Medicine ,Viral tegument ,biology.organism_classification ,Oncomiracidium ,Cell biology ,Infectious Diseases ,Epidermis (zoology) ,Ultrastructure ,Larva ,Insect Science ,Tegument ,Parasitology ,Trematoda ,Epidermis ,Monogenea ,Salmonidae - Abstract
During their different life stages, parasites undergo remarkable morphological, physiological, and behavioral “metamorphoses” to meet the needs of their changing habitats. This is even true for ectoparasites, such as the monogeneans, which typically have a free-swimming larval stage (oncomiracidium) that seeks out and attaches to the external surfaces of fish where they mature. Before any obvious changes occur, there are ultrastructural differences in the oncomiracidium’s outer surface that prepare it for a parasitic existence. The present findings suggest a distinct variation in timing of the switch from oncomiracidia epidermis to the syncytial structure of the adult tegument and so, to date, there are three such categories within the Monogenea: (1) Nuclei of both ciliated cells and interciliary cytoplasm are shed from the surface layer and the epidermis becomes a syncytial layer during the later stages of embryogenesis; (2) nuclei of both ciliated cells and interciliary syncytium remain distinct and the switch occurs later after the oncomiracidia hatch (as in the present study); and (3) the nuclei remain distinct in the ciliated epidermis but those of the interciliary epidermis are lost during embryonic development. Here we describe how the epidermis of the oncomiracidium of Discocotyle sagittata is differentiated into two regions, a ciliated cell layer and an interciliary, syncytial cytoplasm, both of which are nucleated. The interciliary syncytium extends in-between and underneath the ciliated cells and sometimes covers part of their apical surfaces, possibly the start of their shedding process. The presence of membranous whorls and pyknotic nuclei over the surface are indicative of membrane turnover suggesting that the switch in epidermis morphology is already initiated at this stage. The body tegument and associated putative sensory receptors of subadult and adult D. sagittata are similar to those in other monogeneans.
- Published
- 2021
25. A chromatin-associated splicing isoform of OIP5-AS1 acts in cis to regulate the OIP5 oncogene
- Author
-
Michał Wojciech Szcześniak, Elżbieta Wanowska, Izabela Makalowska, and Magdalena Regina Kubiak
- Subjects
Chromosomal Proteins, Non-Histone ,Cyrano ,Cell Cycle Proteins ,Biology ,03 medical and health sciences ,0302 clinical medicine ,lncRNA ,microRNA ,medicine ,Transcriptional regulation ,Humans ,Molecular Biology ,030304 developmental biology ,Cell Proliferation ,0303 health sciences ,OIP5-AS1 ,Alternative splicing ,Cell Biology ,gene expression regulation ,Oncogenes ,Long non-coding RNA ,Chromatin ,Cell biology ,Gene Expression Regulation, Neoplastic ,Cell nucleus ,Alternative Splicing ,medicine.anatomical_structure ,HEK293 Cells ,030220 oncology & carcinogenesis ,RNA splicing ,SMARCA4 ,RNA, Long Noncoding ,Research Article ,Research Paper ,Long noncoding RNA ,OIP5 - Abstract
A large portion of the human genome is transcribed into long noncoding RNAs that can range from 200 nucleotides to several kilobases in length. The number of identified lncRNAs is still growing, but only a handful of them have been functionally characterized. However, it is known that the functions of lncRNAs are closely related to their subcellular localization. Cytoplasmic lncRNAs can regulate mRNA stability, affect translation and act as miRNA sponges, while nuclear-retained long noncoding RNAs have been reported to be involved in transcriptional control, chromosome scaffolding, modulation of alternative splicing and chromatin remodelling. Through these processes, lncRNAs have diverse regulatory roles in cell biology and diseases. OIP5-AS1 (also known as Cyrano), a poorly characterized lncRNA expressed antisense to the OIP5 oncogene, is deregulated in multiple cancers. We showed that one of the OIP5-AS1 splicing forms (ENST00000501665.2) is retained in the cell nucleus where it associates with chromatin, thus narrowing down the spectrum of its possible mechanisms of action. Its knockdown with antisense LNA gapmeRs led to inhibited expression of a sense partner, OIP5, strongly suggesting a functional coupling between OIP5 and ENST00000501665.2. A subsequent bioinformatics analysis followed by RAP-MS and RNA Immunoprecipitation experiments suggested its possible mode of action; in particular, we found that ENST00000501665.2 directly binds to a number of nuclear proteins, including SMARCA4, a component of the SWI/SNF chromatin remodelling complex, whose binding motif is located in the promoter of the OIP5 oncogene.
- Published
- 2021
26. Inhibition of SENP6 restrains cerebral ischemia-reperfusion injury by regulating Annexin-A1 nuclear translocation-associated neuronal apoptosis
- Author
-
Jing Shi, Zhen Zeng, Yin Zhao, Meng Mao, Qian Xia, Zhenzhao Luo, and Xing Li
- Subjects
neuronal apoptosis ,0301 basic medicine ,nuclear translocation ,SENP6 ,Active Transport, Cell Nucleus ,Ischemia ,SUMO protein ,Medicine (miscellaneous) ,Apoptosis ,Mice ,03 medical and health sciences ,Annexin-A1 ,0302 clinical medicine ,medicine ,Animals ,Pharmacology, Toxicology and Pharmaceutics (miscellaneous) ,cerebral ischemia-reperfusion injury ,Annexin A1 ,Ischemic Stroke ,Cell Nucleus ,Neurons ,TUNEL assay ,Chemistry ,medicine.disease ,Cell biology ,Cysteine Endopeptidases ,030104 developmental biology ,Reperfusion Injury ,Phosphorylation ,deSUMOylation ,Reperfusion injury ,030217 neurology & neurosurgery ,Research Paper - Abstract
Rationale: Annexin-A1 (ANXA1) has previously been proposed to play a crucial role in neuronal apoptosis during ischemic stroke injury. Our recent study demonstrated that ANXA1 was modified by SUMOylation, and that this modification was greatly weakened after cerebral ischemia, but its effect on neuronal death and the underlying mechanism have not been fully elucidated. Methods: Mice subjected to middle cerebral artery occlusion were established as the animal model and primary cultured neurons treated with oxygen-glucose deprivation and reperfusion was established as the cell model of ischemic stroke. The Ni2+-NTA agarose affinity pull-down assay was carried out to determine the SUMOylation level of ANXA1. Co-immunoprecipitation assays was utilized to explore the protein interaction. Immunoblot analysis, quantitative real-time PCR, Luciferase reporter assay were performed to identify the regulatory mechanism. LDH release and TUNEL staining was performed to investigate the neuronal cytotoxicity and apoptosis, respectively. Results: In this study, we identified the deSUMOylating enzyme sentrin/SUMO-specific protease 6 (SENP6) as a negative regulator of ANXA1 SUMOylation. Notably, we found that SENP6-mediated deSUMOylation of ANXA1 induced its nuclear translocation and triggered neuronal apoptosis during cerebral ischemic injury. A mechanistic study demonstrated that SENP6-mediated deSUMOylation of ANXA1 promoted TRPM7- and PKC-dependent phosphorylation of ANXA1. Furthermore, blocking the deSUMOylation of ANXA1 mediated by SENP6 inhibited the transcriptional activity of p53, decreased Bid expression, suppressed caspase-3 pathway activation and reduced the apoptosis of primary neurons subjected to oxygen-glucose deprivation and reperfusion. More importantly, SENP6 inhibition by overexpression of a SENP6 catalytic mutant in neurons resulted in significant improvement in neurological function in the mouse model of ischemic stroke. Conclusions: Taken together, the results of this study identified a previously unidentified function of SENP6 in neuronal apoptosis and strongly indicated that SENP6 inhibition may provide therapeutic benefits for cerebral ischemia.
- Published
- 2021
27. YTHDF2 Suppresses Notch Signaling through Post-transcriptional Regulation on Notch1
- Author
-
Byongsun Lee, Seung-Jae Lee, and Jaekyung Shim
- Subjects
Cell ,HES5 ,Notch signaling pathway ,Gene Expression ,Applied Microbiology and Biotechnology ,law.invention ,law ,medicine ,Humans ,Receptor, Notch1 ,HES1 ,Molecular Biology ,Post-transcriptional regulation ,Notch signaling ,Ecology, Evolution, Behavior and Systematics ,Cell Nucleus ,Chemistry ,RNA-Binding Proteins ,Biological Transport ,m6A ,Cell Biology ,Cell biology ,medicine.anatomical_structure ,YTHDF2 ,Apoptosis ,Cytoplasm ,Suppressor ,Heat-Shock Response ,HeLa Cells ,Signal Transduction ,Research Paper ,Developmental Biology - Abstract
YTH domain family 2 (YTHDF2) is an N6-methyladenosine (m6A) binding protein promoting mRNA degradation in various biological processes. Despite its essential roles, the role of YTHDF2 in determining cell fates has not been fully elucidated. Notch signaling plays a vital role in determining cell fates, such as proliferation, differentiation, and apoptosis. We investigated the effect of YTHDF2 on Notch signaling. Our results show that YTHDF2 inhibits Notch signaling by downregulating the Notch1, HES1, and HES5 mRNA levels. Analyzing YTHDF2 deletion mutants indicates that the YTH domain is critical in regulating the Notch signal by directly binding m6A of Notch1 mRNA. Recently, YTHDF2 nuclear translocation was reported under heat shock conditions, but its physiological function is unknown. In our study, the YTH domain is required for YTHDF2 nuclear translocation. In addition, under heat shock stress, the Notch signal was significantly restored due to the increased expression of the Notch1 targets. These results suggest that YTHDF2 in the cytoplasm may act as an intrinsic suppressor in Notch signaling by promoting Notch1 mRNA degradation under normal cellular conditions. Conversely, upon the extracellular stress such as heat shock, YTHDF2 nuclear translocation resulting in reduced Notch1 mRNA decay may contribute to the increasing of Notch intracellular domain (NICD) regulating the survival-related target genes.
- Published
- 2021
28. 25(OH)-Vitamin D alleviates neonatal infectious pneumonia via regulating TGFβ-mediated nuclear translocation mechanism of YAP/TAZ
- Author
-
Qian Liu, Qi Sun, Lina Qiao, Yiwen Gao, and Yi Yuan
- Subjects
Male ,medicine.medical_specialty ,Lipopolysaccharide ,neonatal infectious pneumonia (nip) ,Immunocytochemistry ,Bioengineering ,Applied Microbiology and Biotechnology ,Peripheral blood mononuclear cell ,Calcitriol receptor ,25-oh-vd ,chemistry.chemical_compound ,Transforming Growth Factor beta ,Internal medicine ,medicine ,Vitamin D and neurology ,Humans ,Vitamin D ,Cell Nucleus ,Infant, Newborn ,YAP-Signaling Proteins ,General Medicine ,Pneumonia ,Vitamins ,deficiency ,medicine.disease ,Protein Transport ,Endocrinology ,chemistry ,Case-Control Studies ,yap/taz ,NIP ,Female ,inflammatory indicators ,Acyltransferases ,TP248.13-248.65 ,Transforming growth factor ,Research Article ,Research Paper ,anti-inflammatory mechanism ,Biotechnology - Abstract
Neonatal infectious pneumonia (NIP) is a common infectious disease that develops in the neonatal period. The purpose of our study was to explore the potential roles of 25(OH)-Vitamin D (25-OH-VD) and its anti-inflammatory mechanism in NIP. The results showed that serum 25-OH-VD level was negatively correlated with the severity of NIP, whereas Spearman’s correlation analysis showed a significant positive correlation between the severity of NIP and the levels of pneumonia markers procalcitonin (PCT) and interleukin-6 (IL-6). The expression of vitamin D receptor (VDR) was down-regulated, while the transforming growth factor β (TGFβ), nuclear YAP, and TAZ were up-regulated in the peripheral blood mononuclear cells (PBMCs) of neonates with severe pneumonia. Neonates with 25-OH-VD deficiency were associated with an increased risk of NIP. In BEAS-2B cells, down-regulation of nuclear YAP and TAZ was found in the lipopolysaccharide (LPS) + VD group relative to the LPS-induced group. Additionally, positive rate of nuclear YAP, as detected by immunocytochemistry (ICC), and the nuclear translocation of nuclear YAP/TAZ by IFA in the LPS+VD group showed an intermediate level between that of the control and LPS-induced groups. Furthermore, the expressions of VDR and CYP27B1 were significantly increased in the LPS+VD group as compared to those in the LPS-induced group. The anti-inflammatory mechanism in NIP was achieved due to the 25-OH-VD mediating TGFβ/YAP/TAZ pathway, which suggested that using 25-OH-VD might be a potential strategy for NIP treatment.
- Published
- 2021
29. TMT-based quantitative proteomics analysis reveals the attenuated replication mechanism of Newcastle disease virus caused by nuclear localization signal mutation in viral matrix protein
- Author
-
Mengmeng Ni, Yong Ruan, Jiaqi Chen, Jiafu Zhao, Zhiqiang Duan, Yifan Han, Lei Zhou, Xinqin Ji, and Chao Yuan
- Subjects
Proteomics ,Microbiology (medical) ,quantitative proteomics ,Newcastle Disease ,viruses ,Nuclear Localization Signals ,Immunology ,matrix protein ,Infectious and parasitic diseases ,RC109-216 ,Virus Replication ,Microbiology ,Virus ,Cell Line ,Viral Matrix Proteins ,03 medical and health sciences ,nucleocytoplasmic trafficking ,Transcription (biology) ,Cricetinae ,Protein biosynthesis ,tmt ,Animals ,NLS ,030304 developmental biology ,Cell Nucleus ,0303 health sciences ,Viral matrix protein ,Host Microbial Interactions ,biology ,030306 microbiology ,RNA virus ,newcastle disease virus ,biology.organism_classification ,nuclear localization signal ,Cell biology ,Infectious Diseases ,Viral replication ,Mutation ,Parasitology ,Nuclear localization sequence ,Research Article ,Research Paper - Abstract
Nuclear localization of cytoplasmic RNA virus proteins mediated by intrinsic nuclear localization signal (NLS) plays essential roles in successful virus replication. We previously reported that NLS mutation in the matrix (M) protein obviously attenuates the replication and pathogenicity of Newcastle disease virus (NDV), but the attenuated replication mechanism remains unclear. In this study, we showed that M/NLS mutation not only disrupted M’s nucleocytoplasmic trafficking characteristic but also impaired viral RNA synthesis and transcription. Using TMT-based quantitative proteomics analysis of BSR-T7/5 cells infected with the parental NDV rSS1GFP and the mutant NDV rSS1GFP-M/NLSm harboring M/NLS mutation, we found that rSS1GFP infection stimulated much greater quantities and more expression changes of differentially expressed proteins involved in host cell transcription, ribosomal structure, posttranslational modification, and intracellular trafficking than rSS1GFP-M/NLSm infection. Further in-depth analysis revealed that the dominant nuclear accumulation of M protein inhibited host cell transcription, RNA processing and modification, protein synthesis, posttranscriptional modification and transport; and this kind of inhibition could be weakened when most of M protein was confined outside the nucleus. More importantly, we found that the function of M protein in the cytoplasm effected the inhibition of TIFA expression in a dose-dependent manner, and promoted NDV replication by down-regulating TIFA/TRAF6/NF-κB-mediated production of cytokines. It was the first report about the involvement of M protein in NDV immune evasion. Taken together, our findings demonstrate that NDV replication is closely related to the nucleocytoplasmic trafficking of M protein, which accelerates our understanding of the molecular functions of NDV M protein.
- Published
- 2020
30. Cell differentiation and aging accompanied by depletion of the ACE2 protein
- Author
-
Soňa Legartová, Orazio Angelo Arcidiacono, Jana Krejčí, and Eva Bártová
- Subjects
Male ,Aging ,medicine.medical_specialty ,Cell type ,embryonic heart ,lung cancer cells ,Cellular differentiation ,ACE2 ,Renin-Angiotensin System ,Mice ,Sex Factors ,Downregulation and upregulation ,Cell surface receptor ,Internal medicine ,Renin–angiotensin system ,medicine ,Animals ,Humans ,Pandemics ,SARS-CoV-2 ,Chemistry ,Age Factors ,COVID-19 ,Cell Differentiation ,Embryo ,Cell Biology ,Pathophysiology ,Cell nucleus ,HEK293 Cells ,Endocrinology ,medicine.anatomical_structure ,renin ,Gene Expression Regulation ,A549 Cells ,human kidney embryonic cells ,Female ,Angiotensin-Converting Enzyme 2 ,HT29 Cells ,hormones, hormone substitutes, and hormone antagonists ,Research Paper - Abstract
ACE2 was observed as the cell surface receptor of the SARS-CoV-2 virus. Interestingly, we also found ACE2 positivity inside the cell nucleus. The ACE2 levels changed during cell differentiation and aging and varied in distinct cell types. We observed ACE2 depletion in the aortas of aging female mice, similarly, the aging caused ACE2 decrease in the kidneys. Compared with that in the heart, brain and kidneys, the ACE2 level was the lowest in the mouse lungs. In mice exposed to nicotine, ACE2 was not changed in olfactory bulbs but in the lungs, ACE2 was upregulated in females and downregulated in males. These observations indicate the distinct gender-dependent properties of ACE2. Differentiation into enterocytes, and cardiomyocytes, caused ACE2 depletion. The cardiomyogenesis was accompanied by renin upregulation, delayed in HDAC1-depleted cells. In contrast, vitamin D2 decreased the renin level while ACE2 was upregulated. Together, the ACE2 level is high in non-differentiated cells. This protein is more abundant in the tissues of mouse embryos and young mice in comparison with older animals. Mostly, downregulation of ACE2 is accompanied by renin upregulation. Thus, the pathophysiology of COVID-19 disease should be further studied not only by considering the ACE2 level but also the whole renin-angiotensin system.
- Published
- 2020
31. Large-scale analysis of BAP1 expression reveals novel associations with clinical and molecular features of malignant pleural mesothelioma
- Author
-
William G. Richards, Assunta De Rienzo, Samuel Freyaldenhoven, Roderick V. Jensen, Claire V. Meyerovitz, Michela E. Oster, Corinne E. Gustafson, Beow Y. Yeap, David T. Severson, Yin P Hung, Nhien T. Dao, Lucian R. Chirieac, Raphael Bueno, and Biological Sciences
- Subjects
Adult ,Male ,Pathology ,medicine.medical_specialty ,Epithelial-Mesenchymal Transition ,Tumor suppressor gene ,Adolescent ,Pleural Neoplasms ,DNA Mutational Analysis ,Vimentin ,Biology ,epithelial‐to‐mesenchymal transition ,Pathology and Forensic Medicine ,Young Adult ,medicine ,Biomarkers, Tumor ,Humans ,BAP1 ,Mesothelioma ,tumor suppressor gene ,prognostic biomarker ,Aged ,Aged, 80 and over ,Cell Nucleus ,Original Paper ,Tumor Suppressor Proteins ,Mesothelioma, Malignant ,intra‐tumor heterogeneity ,intra-tumor heterogeneity ,Histology ,Middle Aged ,medicine.disease ,Prognosis ,Immunohistochemistry ,Original Papers ,Staining ,Tumor progression ,mesothelioma ,Mutation ,immunohistochemistry ,biology.protein ,gene expression ,Female ,epithelial-to-mesenchymal transition ,Ubiquitin Thiolesterase - Abstract
BRCA1-associated protein-1(BAP1) expression is commonly lost in several tumors including malignant pleural mesothelioma (MPM). Presence or absence of immunohistochemical BAP1 nuclear staining in tumor cells is currently used for differential diagnosis of MPM. In this study, a large cohort of 596 MPM tumors with available clinical data was analyzed to examine associations of BAP1 staining pattern with clinical and molecular features that may reflect the impact ofBAP1mutation on MPM biology. Cases were classified according to the BAP1 staining pattern of tumor cells. Exome and RNA-sequencing data were available for subsets of cases. Levels of mRNA encoding claudin 15 (CLDN15) and vimentin (VIM) were determined using RT-qPCR on 483 cases to estimate the relative proportions of epithelial-like and mesenchymal-like components in each tumor. Four BAP1 staining patterns were observed: single-pattern nuclear staining (36%), single-pattern cytoplasmic staining (25%), single-pattern absent staining (12%), and combinations of these staining patterns (27%). This study confirmed prior reports that nuclear BAP1 is more frequently associated with wild-typeBAP1and sarcomatoid histology. However, no associations between BAP1 staining pattern(s) and mutations in specific protein domains and/or mutation type were observed. BAP1 staining patterns were significantly associated (p < 0.001) withBAP1gene expression, MPM histologic subtypes, molecular clusters, and markers of epithelial-to-mesenchymal transition. Frequent observation of combinations of BAP1 staining patterns in MPM tumors indicated intra-tumoral heterogeneity ofBAP1status. Cytoplasmic BAP1 staining was identified as a putative indicator of favorable prognosis in non-epithelioid MPM. In conclusion, novel significant associations among different BAP1 staining patterns and subgroups of MPM tumors were observed, suggesting that the role ofBAP1in tumor progression may be more complex than its presumed tumor suppressor function. Cytoplasmic staining was identified as a putative indicator of favorable prognosis in non-epithelioid MPM, potentially addressing a critical need in clinical decision-making in this disease. (c) 2020 The Authors.The Journal of Pathologypublished by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland. National Cancer InstituteUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [NCI 2 R01 CA120528-11A1]; International Mesothelioma Program at Brigham and Women's Hospital; NCI Cancer Center Support GrantUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [NIH 5 P30 CA06516] This work was supported by grants to RB from the National Cancer Institute (NCI 2 R01 CA120528-11A1) and the International Mesothelioma Program at Brigham and Women's Hospital. The study sponsors played no role in the study design, collection, analysis, interpretation of data, writing of the report, or decision to submit the paper for publication. We thank the Dana-Farber/Harvard Cancer Center in Boston, MA, for the use of the Specialized Histopathology Core, which provided histology and immunohistochemistry service. Dana-Farber/Harvard Cancer Center is supported in part by an NCI Cancer Center Support Grant #NIH 5 P30 CA06516.
- Published
- 2020
32. Type 2 diabetes causes skeletal muscle atrophy but does not impair resistance training‐mediated myonuclear accretion and muscle mass gain in rats
- Author
-
Koji Sato, Kohei Kido, Satoshi Fujita, and Satoru Ato
- Subjects
Male ,endocrine system diseases ,Physiology ,medicine.medical_treatment ,Muscle Fibers, Skeletal ,Type 2 diabetes ,030204 cardiovascular system & hematology ,Muscle hypertrophy ,0302 clinical medicine ,RESEARCH PAPERS ,Insulin ,Phosphorylation ,Nutrition and Dietetics ,Organ Size ,General Medicine ,MUSCLE ,Adaptation, Physiological ,Muscle atrophy ,Muscular Atrophy ,medicine.anatomical_structure ,Adipose Tissue ,type 2 diabetes ,medicine.symptom ,Research Paper ,medicine.medical_specialty ,Rats, Inbred OLETF ,Mechanistic Target of Rapamycin Complex 1 ,03 medical and health sciences ,Gastrocnemius muscle ,Physiology (medical) ,Diabetes mellitus ,Internal medicine ,medicine ,Animals ,Hypoglycemic Agents ,muscle hypertrophy ,Muscle, Skeletal ,Cell Nucleus ,business.industry ,nutritional and metabolic diseases ,Skeletal muscle ,Type 2 Diabetes Mellitus ,Resistance Training ,Editor's Pick ,medicine.disease ,Rats ,Endocrinology ,Diabetes Mellitus, Type 2 ,business ,030217 neurology & neurosurgery - Abstract
New findings What is the central question of this study? Type 2 diabetes mellitus (T2DM) causes skeletal muscle atrophy; does it affect resistance training (RT)-mediated molecular adaptations and subsequent muscle hypertrophy? What is the main finding and its importance? Although skeletal muscle mass and regulation were not preserved under conditions of T2DM, the response of RT-induced skeletal muscle hypertrophy was not impaired in T2DM rat skeletal muscle. These findings suggest that the capacity of RT-mediated muscle mass gain is not diminished in the T2DM condition. Abstract Type 2 diabetes mellitus (T2DM) is known to cause skeletal muscle atrophy. However, it is not known whether T2DM affects resistance training (RT)-mediated molecular adaptations and subsequent muscle hypertrophy. Therefore, we investigated the effect of T2DM on response of skeletal muscle hypertrophy to chronic RT using a rat resistance exercise mimetic model. T2DM and healthy control rats were subjected to 18 bouts (3 times per week) of chronic RT on unilateral lower legs. RT significantly increased gastrocnemius muscle mass and myonuclei in both T2DM and healthy control rats to the same extent, even though T2DM caused muscle atrophy in the resting condition. Further, T2DM significantly reduced mechanistic target of rapamycin complex 1 (mTORC1) activity (phosphorylation of p70S6KThr389 and 4E-BP1Thr37/46 ) to insulin stimulation and the number of myonuclei in the untrained basal condition, but RT-mediated adaptations were not affected by T2DM. These findings suggested that although the skeletal muscle mass and regulation were not preserved under basal conditions of T2DM, the response of RT-induced skeletal muscle hypertrophy was not impaired in T2DM rat skeletal muscle.
- Published
- 2019
33. F‐box protein FBXO16 functions as a tumor suppressor by attenuating nuclear β‐catenin function
- Author
-
Praveen Kumar Shetty, Biswanath Maity, Srikanth Rapole, Ramanamurthy Boppana, Debasish Paul, Rajesh Kumar Manne, US Dinesh, Sehbanul Islam, Manas Kumar Santra, and Sunil K. Malonia
- Subjects
0301 basic medicine ,proteasomal degradation ,Epithelial-Mesenchymal Transition ,tumor suppressor ,F-box protein ,Pathology and Forensic Medicine ,law.invention ,03 medical and health sciences ,0302 clinical medicine ,Ubiquitin ,law ,Cell Line, Tumor ,medicine ,Humans ,Genes, Tumor Suppressor ,Wnt Signaling Pathway ,beta Catenin ,Cell Nucleus ,Original Paper ,biology ,Chemistry ,F-Box Proteins ,Wnt signaling pathway ,Nuclear Proteins ,Cancer ,medicine.disease ,Original Papers ,Wnt signaling ,Gene Expression Regulation, Neoplastic ,030104 developmental biology ,Proteasome ,FBXO16 ,030220 oncology & carcinogenesis ,Catenin ,Cancer cell ,biology.protein ,Cancer research ,β‐catenin ,Suppressor - Abstract
Aberrant activation of β‐catenin has been implicated in a variety of human diseases, including cancer. In spite of significant progress, the regulation of active Wnt/β‐catenin‐signaling pathways is still poorly understood. In this study, we show that F‐box protein 16 (FBXO16) is a putative tumor suppressor. It is a component of the SCF (SKP1‐Cullin1‐F‐box protein) complex, which targets the nuclear β‐catenin protein to facilitate proteasomal degradation through the 26S proteasome. FBXO16 interacts physically with the C‐terminal domain of β‐catenin and promotes its lysine 48‐linked polyubiquitination. In addition, it inhibits epithelial‐to‐mesenchymal transition (EMT) by attenuating the level of β‐catenin. Therefore, depletion of FBXO16 leads to increased levels of β‐catenin, which then promotes cell invasion, tumor growth, and EMT of cancer cells. Furthermore, FBXO16 and β‐catenin share an inverse correlation of cellular expression in clinical breast cancer patient samples. In summary, we propose that FBXO16 functions as a putative tumor suppressor by forming an SCFFBXO16 complex that targets nuclear β‐catenin in a unique manner for ubiquitination and subsequent proteasomal degradation to prevent malignancy. This work suggests a novel therapeutic strategy against human cancers related to aberrant β‐catenin activation. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
- Published
- 2019
34. Proposal for determining absolute biological effectiveness of boron neutron capture therapy—the effect of 10B(n,α)7Li dose can be predicted from the nucleocytoplasmic ratio or the cell size
- Author
-
Yuki Tamari, Minoru Suzuki, Hiroki Tanaka, Shin-ichiro Masunaga, Koji Ono, and Tsubasa Watanabe
- Subjects
inorganic chemicals ,Health, Toxicology and Mutagenesis ,Boron Neutron Capture Therapy ,Lithium ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Radiation sensitivity ,Cell Line, Tumor ,Regular Paper ,medicine ,Relative biological effectiveness ,Animals ,Humans ,Radiology, Nuclear Medicine and imaging ,Radiosensitivity ,Biology ,Boron ,Cell Nucleus ,Neutrons ,Radiation ,Chemistry ,business.industry ,Melanoma ,Dose-Response Relationship, Radiation ,medicine.disease ,BPA ,Neutron temperature ,D0 ,cell size ,Neutron capture ,Dose–response relationship ,030220 oncology & carcinogenesis ,BSH, N/C ratio ,Cell survival curve ,Nuclear medicine ,business ,Relative Biological Effectiveness ,boron-neutron dose - Abstract
The relationship between the radiation dose delivered to a tumor and its effect is not completely predictable. Uncertainty in the estimation of the boron concentration in a tumor, variation in the radiation sensitivity of the tumor cells, and the complexity of the interactions between the four types of radiation comprising the boron neutron capture therapy (BNCT) dose contribute to this uncertainty. We reanalyzed the data of our previous papers to investigate the variation in radiosensitivity of tumor cells to the 10B(n,α)7Li dose: the dose generated by the reaction of thermal neutrons and 10B, hereafter the ‘boron-neutron dose’. The radiosensitivities of five tumors (EL4, SAS/neo, SAS/mp53, SCCVII and B16-BL6 melanoma) were examined. For the combination of p-boron-L-phenylalanine (BPA: C9H12BNO4) with neutron irradiation, D0, the cell survival curve for the boron-neutron dose was the smallest for the SAS/neo, followed by the EL4, SAS/mp53, SCCVII and B16-BL6 melanoma, in that order. For the combination of mercaptoundecahydrododecaborate (BSH: Na2B12H11SH) with neutron irradiation, D0 was the smallest for the EL4, followed by the SAS/neo, B16–BL6melanoma, SAS/mp53 and SCCVII, in that order. The relationships between these D0 values and the nucleocytoplasmic ratios (Xncs) or cell size indices (Xcs) obtained by histopathological microslide image were as follows: (D0 = 0.1341Xnc–1.586, R2 = 0.9721) for all tumor types with BPA-BNCT, and D0 = 0.0122Xcs–0.1319 (R2 = 0.9795) for four tumor types (all except the B16-BL6 melanoma) with BSH-BNCT. Based on these results, we proposed a new biologically equivalent effectiveness factor: the absolute biological effectiveness (ABE) factor. The ABE factor is Gy/D0. Thus, the ABE dose is the physical dose multiplied by the ABE factor, and refers to the dose needed to decrease the cell survival rate to e–ABE dose/Gy.
- Published
- 2018
35. The E3 ligase RFWD3 stabilizes ORC in a p53-dependent manner
- Author
-
Sumanprava Giri, Susan Wopat, Supriya G. Prasanth, Dazhen Liu, Arindam Chakraborty, Yating Wang, Yo-Chuen Lin, Rosaline Y.C. Hsu, and Kannanganattu V. Prasanth
- Subjects
DNA Replication ,0301 basic medicine ,DNA damage ,Ubiquitin-Protein Ligases ,Origin Recognition Complex ,Cell Line ,03 medical and health sciences ,0302 clinical medicine ,Ubiquitin ,Cell Line, Tumor ,Humans ,Molecular Biology ,Cell Nucleus ,biology ,Binding protein ,Ubiquitination ,DNA replication ,Wild type ,Cell Biology ,Chromatin ,Ubiquitin ligase ,Proliferating cell nuclear antigen ,Cell biology ,HEK293 Cells ,030104 developmental biology ,030220 oncology & carcinogenesis ,Microtubule Proteins ,biology.protein ,Origin recognition complex ,Tumor Suppressor Protein p53 ,Research Paper ,DNA Damage ,Protein Binding ,Developmental Biology - Abstract
RFWD3 is an E3 ubiquitin ligase that plays important roles in DNA damage response and DNA replication. We have previously demonstrated that the stabilization of RFWD3 by PCNA at the replication fork enables ubiquitination of the single-stranded binding protein, RPA and its subsequent degradation for replication progression. Here, we report that RFWD3 associates with the Origin Recognition Complex (ORC) and ORC-Associated (ORCA/LRWD1), components of the pre-replicative complex required for the initiation of DNA replication. Overexpression of ORC/ORCA leads to the stabilization of RFWD3. Interestingly, RFWD3 seems to stabilize ORC/ORCA in cells expressing wild type p53, as the depletion of RFWD3 reduces the levels of ORC/ORCA. Further, the catalytic activity of RFWD3 is required for the stabilization of ORC. Our results indicate that the RFWD3 promotes the stability of ORC, enabling efficient pre-RC assembly.
- Published
- 2020
36. BMP6/TAZ-Hippo signaling modulates angiogenesis and endothelial cell response to VEGF
- Author
-
Johanna P. Laakkonen, Henri Niskanen, Lari Holappa, Jari P. Lappalainen, Seppo Ylä-Herttuala, H. H. Pulkkinen, Minna U. Kaikkonen, Mustafa Beter, M. Kiema, Annakaisa Tirronen, and Anu Toropainen
- Subjects
0301 basic medicine ,Male ,Vascular Endothelial Growth Factor A ,Cancer Research ,animal structures ,Physiology ,Angiogenesis ,Bone Morphogenetic Protein 6 ,Swine ,Clinical Biochemistry ,Notch signaling pathway ,BMP2 ,Bone Morphogenetic Protein 2 ,Neovascularization, Physiologic ,BMP6 ,Protein Serine-Threonine Kinases ,Bone morphogenetic protein ,Models, Biological ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Animals ,Humans ,Hippo Signaling Pathway ,Adaptor Proteins, Signal Transducing ,Cell Nucleus ,Hippo signaling pathway ,Original Paper ,Base Sequence ,Endothelial Cells ,Kinase insert domain receptor ,VEGF ,Vascular Endothelial Growth Factor Receptor-2 ,Cell Hypoxia ,3. Good health ,Cell biology ,Vascular endothelial growth factor ,Mice, Inbred C57BL ,Protein Transport ,030104 developmental biology ,VEGFR2 ,chemistry ,Hippo signaling ,030220 oncology & carcinogenesis ,Signal transduction ,Signal Transduction - Abstract
The BMP/TGFβ-Smad, Notch and VEGF signaling guides formation of endothelial tip and stalk cells. However, the crosstalk of bone morphogenetic proteins (BMPs) and vascular endothelial growth factor receptor 2 (VEGFR2) signaling has remained largely unknown. We demonstrate that BMP family members regulate VEGFR2 and Notch signaling, and act via TAZ-Hippo signaling pathway. BMPs were found to be regulated after VEGF gene transfer in C57/Bl6 mice and in a porcine myocardial ischemia model. BMPs 2/4/6 were identified as endothelium-specific targets of VEGF. BMP2 modulated VEGF-mediated endothelial sprouting via Delta like Canonical Notch Ligand 4 (DLL4). BMP6 modulated VEGF signaling by regulating VEGFR2 expression and acted via Hippo signaling effector TAZ, known to regulate cell survival/proliferation, and to be dysregulated in cancer. In a matrigel plug assay in nude mice BMP6 was further demonstrated to induce angiogenesis. BMP6 is the first member of BMP family found to directly regulate both Hippo signaling and neovessel formation. It may thus serve as a target in pro/anti-angiogenic therapies. Electronic supplementary material The online version of this article (10.1007/s10456-020-09748-4) contains supplementary material, which is available to authorized users.
- Published
- 2020
37. LINC-complex mediated positioning of the vegetative nucleus is involved in calcium and ROS signaling in Arabidopsis pollen tubes
- Author
-
Norman Reid Groves, Andrew B. Kirkpatrick, Morgan Moser, and Iris Meier
- Subjects
lcsh:QH426-470 ,LINC complex ,Arabidopsis ,Pollen Tube ,03 medical and health sciences ,medicine ,otorhinolaryngologic diseases ,Inner membrane ,Pollen tube tip ,Nuclear Matrix ,lcsh:QH573-671 ,Cytoskeleton ,030304 developmental biology ,reactive oxygen species ,Cell Nucleus ,0303 health sciences ,biology ,lcsh:Cytology ,Chemistry ,Arabidopsis Proteins ,030302 biochemistry & molecular biology ,nuclear calcium ,food and beverages ,Cell Biology ,nuclear envelope ,biology.organism_classification ,Cell biology ,pollen tube termination ,lcsh:Genetics ,Pollen tube reception ,medicine.anatomical_structure ,Male fertility ,Pollen tube ,Calcium ,Nucleus ,Research Article ,Research Paper ,Signal Transduction - Abstract
Nuclear movement and positioning play a role in developmental processes throughout life. Nuclear movement and positioning are mediated primarily by linker of nucleoskeleton and cytoskeleton (LINC) complexes. LINC complexes are comprised of the inner nuclear membrane SUN proteins and the outer nuclear membrane (ONM) KASH proteins. In Arabidopsis pollen tubes, the vegetative nucleus (VN) maintains a fixed distance from the pollen tube tip during growth, and the VN precedes the sperm cells (SCs). In pollen tubes of wit12 and wifi, mutants deficient in the ONM component of a plant LINC complex, the SCs precede the VN during pollen tube growth and the fixed VN distance from the tip is lost. Subsequently, pollen tubes frequently fail to burst upon reception. In this study, we sought to determine if the pollen tube reception defect observed in wit12 and wifi is due to decreased sensitivity to reactive oxygen species (ROS). Here, we show that wit12 and wifi are hyposensitive to exogenous H2O2, and that this hyposensitivity is correlated with decreased proximity of the VN to the pollen tube tip. Additionally, we report the first instance of nuclear Ca2+ peaks in growing pollen tubes, which are disrupted in the wit12 mutant. In the wit12 mutant, nuclear Ca2+ peaks are reduced in response to exogenous ROS, but these peaks are not correlated with pollen tube burst. This study finds that VN proximity to the pollen tube tip is required for both response to exogenous ROS, as well as internal nuclear Ca2+ fluctuations.
- Published
- 2020
38. Cytoplasmic PCNA is located in the actin belt and involved in osteoclast differentiation
- Author
-
Huan Tian Zhang, Xiaohui Liu, Xuejuan Gao, Jiake Xu, Yong Dai, Kezhi Chen, Langxia Liu, Donge Tang, and Zhipeng Li
- Subjects
Male ,musculoskeletal diseases ,Cytoplasm ,Aging ,Primary Cell Culture ,Osteoclasts ,Bone resorption ,Mice ,Multinucleate ,Osteoclast ,Proliferating Cell Nuclear Antigen ,medicine ,Animals ,Humans ,PCNA ,Nuclear protein ,Actin ,Cell Nucleus ,Gene knockdown ,biology ,Chemistry ,RANK Ligand ,Cell Differentiation ,differentiation ,Cell Biology ,Actins ,Cell biology ,Proliferating cell nuclear antigen ,Actin Cytoskeleton ,Disease Models, Animal ,RAW 264.7 Cells ,medicine.anatomical_structure ,Gene Knockdown Techniques ,osteoclast ,biology.protein ,Osteoporosis ,actin belt ,Research Paper - Abstract
Osteoporosis (OP) is an age-related osteolytic disease and characterized by low bone mass and more prone to fracture due to active osteoclasts. Proliferating cell nuclear antigen (PCNA) has been long identified as a nuclear protein playing critical roles in the regulation of DNA replication and repair. Recently, a few studies have demonstrated the cytoplasmic localization of PCNA and its function associated with apoptosis in neutrophil and neuroblastoma cells. However, the involvement of PCNA, including the cytoplasmic PCNA, in the osteoclast differentiation remains unclear. In the present study, we show that PCNA is translocated from nucleus to cytoplasm during the RANKL-induced osteoclast differentiation, and localized in the actin belt of mature osteoclast. Knockdown of PCNA significantly affected the integrity of actin belt, the formation of multinucleated osteoclasts, the expression of osteoclast-specific genes, and the in vitro bone resorption. Interactomic study has revealed β-actin as the major interacting partner of the cytoplasmic PCNA, suggesting that cytoplasmic PCNA might play a critical role in the differentiation of osteoclast through regulation of actin-cytoskeleton remodeling. Taken together, our results demonstrate the critical role of cytoplasmic PCNA during the process of osteoclast differentiation, and provided a potential therapeutic target for treatment of osteoclast-related bone diseases.
- Published
- 2020
39. Relationship between thyroid tumor radiosensitivity and nuclear localization of DNA-dependent protein kinase catalytic subunit
- Author
-
Masahiro Nakashima, Kiyoto Ashizawa, Ichiro Horie, Ryota Otsubo, Makoto Ihara, Katsuya Matsuda, Takashi Kudo, and Kazuko Shichijo
- Subjects
Cytoplasm ,DNA End-Joining Repair ,DNA Repair ,thyroid tumor ,Health, Toxicology and Mutagenesis ,DNA-Activated Protein Kinase ,Radiation Tolerance ,DNA-Dependent Protein Kinase Catalytic Subunit ,Iodine Radioisotopes ,03 medical and health sciences ,0302 clinical medicine ,Protein Domains ,Catalytic Domain ,Cell Line, Tumor ,medicine ,Regular Paper ,Humans ,Radiology, Nuclear Medicine and imaging ,DNA Breaks, Double-Stranded ,Radiosensitivity ,Thyroid Neoplasms ,Thyroid cancer ,DNA-PKcs ,030304 developmental biology ,Cell Nucleus ,0303 health sciences ,Radiation ,biology ,Chemistry ,Thyroid ,medicine.disease ,Molecular biology ,Immunohistochemistry ,enzymes and coenzymes (carbohydrates) ,medicine.anatomical_structure ,Microscopy, Fluorescence ,Cell culture ,immunohistochemical staining ,030220 oncology & carcinogenesis ,Cancer cell ,biology.protein ,radiosensitivity prediction ,AcademicSubjects/SCI00960 ,Antibody ,biological phenomena, cell phenomena, and immunity - Abstract
Thyroid tumors are the most common types of endocrine malignancies and are commonly treated with radioactive iodine (RAI) to destroy remaining cancer cells following surgical intervention. We previously reported that the expression levels of double-stranded DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which plays a key role in non-homologous end joining, are correlated with the radiosensitivity of cancer cells. Specifically, cells expressing high levels of DNA-PKcs exhibited radiation resistance, whereas cells expressing low levels were sensitive to radiation treatment. In this study, we observed full-length native DNA-PKcs (460 kDa) in radiation-resistant FRO and KTC-2 cells through western blot analysis using an antibody against the C-terminus of DNA-PKcs. In contrast, cleaved DNA-PKcs (175 kDa) were observed in radiation-sensitive TPC-1 and KTC-1 cells. Almost equal amounts of DNA-PKcs were observed in moderately radiation-sensitive WRO cells. We also describe a simple method for the prediction of radiation therapy efficacy in individual cases of thyroid cancers based on staining for DNA-PKcs in human cancer cell lines. Immunofluorescent staining showed that native DNA-PKcs was localized largely in the cytoplasm and only rarely localized in the nuclei of radiation-resistant thyroid cancer cells, whereas in radiation-sensitive cancer cells a 175-kDa cleaved C-terminal fragment of DNA-PKcs was localized mainly inside the nuclei. Therefore, DNA-PKcs moved to the nucleus after γ-ray irradiation. Our results suggest a new method for classifying human thyroid tumors based on their cellular distribution patterns of DNA-PKcs in combination with their radiosensitivity., Journal of radiation research, 61(4), pp.511-516; 2020
- Published
- 2020
40. Epothilone B prevents lipopolysaccharide-induced inflammatory osteolysis through suppressing osteoclastogenesis via STAT3 signaling pathway
- Author
-
Fei Luo, Jiulin Tan, Wenhui Hu, Yueqi Chen, Zhiyong Hou, Fanchun Zeng, Yong Tang, Yiran Wang, Jianzhong Xu, Shiwu Dong, Zhansong Tian, Junxian Hu, and Qijie Dai
- Subjects
Lipopolysaccharides ,STAT3 Transcription Factor ,Aging ,Osteolysis ,Primary Cell Culture ,Osteoclasts ,Apoptosis ,Osteoclast fusion ,Bone resorption ,Stat3 Signaling Pathway ,STAT3 ,Mice ,Cytosol ,Osteoclast ,Osteogenesis ,inflammatory osteolysis ,medicine ,Animals ,Humans ,Femur ,Epothilone B (Epo B) ,Protein kinase B ,PI3K/AKT/mTOR pathway ,osteoclastogenesis ,Cell Nucleus ,biology ,NFATC Transcription Factors ,Chemistry ,RANK Ligand ,NF-kappa B ,Transcription Factor RelA ,Cell Differentiation ,Cell Biology ,X-Ray Microtomography ,medicine.disease ,Cell biology ,Disease Models, Animal ,medicine.anatomical_structure ,RAW 264.7 Cells ,RANKL ,Epothilones ,biology.protein ,Female ,lipopolysaccharide (LPS) ,Research Paper ,Signal Transduction - Abstract
Inflammatory osteolysis is a common osteolytic specificity that occurs during infectious orthopaedic surgery and is characterized by an imbalance in bone homeostasis due to excessive osteoclast bone resorption activity. Epothilone B (Epo B) induced α-tubulin polymerization and enhanced microtubule stability, which also played an essential role in anti-inflammatory effect on the regulation of many diseases. However, its effects on skeletal system have rarely been investigated. Our study demonstrated that Epo B inhibited osteoclastogenesis in vitro and prevented inflammatory osteolysis in vivo. Further analysis showed that Epo B also markedly induced mature osteoclasts apoptosis during osteoclastogenesis. Mechanistically, Epo B directly suppressed osteoclastogenesis by the inhibitory regulation of the phosphorylation and activation of PI3K/Akt/STAT3 signaling directly, and the suppressive regulation of the CD9/gp130/STAT3 signaling pathway indirectly. The negative regulatory effect on STAT3 signaling further restrained the translocation of NF-κB p65 and NFATc1 from the cytosol to the nuclei during RANKL stimulation. Additionally, the expression of osteoclast specific genes was also significantly attenuated during osteoclast fusion and differentiation. Taken together, these findings illustrated that Epo B protected against LPS-induced bone destruction through inhibiting osteoclastogenesis via regulating the STAT3 dependent signaling pathway.
- Published
- 2020
41. FAM84B, amplified in pancreatic ductal adenocarcinoma, promotes tumorigenesis through the Wnt/β-catenin pathway
- Author
-
Dejun Yang, Zunqi Hu, Qing You, Qingping Cai, Ronglin Yan, Yu Zhang, Jiapeng Xu, Xin Zhang, and Hongbing Fu
- Subjects
Male ,Aging ,endocrine system diseases ,Carcinogenesis ,Survivin ,Gene Expression ,Apoptosis ,medicine.disease_cause ,Deoxycytidine ,Mice ,Wnt Signaling Pathway ,beta Catenin ,Wnt/β-catenin ,Gene knockdown ,education.field_of_study ,Chemistry ,gemcitabine ,Wnt signaling pathway ,glycolysis ,Middle Aged ,Mitochondria ,Neoplasm Proteins ,Survival Rate ,Gene Knockdown Techniques ,Female ,Research Paper ,Carcinoma, Pancreatic Ductal ,Antimetabolites, Antineoplastic ,proliferation ,Lactate dehydrogenase A ,Proto-Oncogene Proteins c-myc ,Cell Line, Tumor ,medicine ,Animals ,Humans ,education ,Cell Proliferation ,Cell Nucleus ,Cell growth ,Gene Amplification ,Membrane Proteins ,Cell Biology ,Pancreatic Neoplasms ,Anaerobic glycolysis ,Catenin ,Cancer research ,Lactate Dehydrogenase 5 ,Neoplasm Transplantation - Abstract
Altered expression of family with sequence similarity 84, member B (FAM84B) has been found in various human cancers. However, the expression and function of FAM84B in pancreatic ductal adenocarcinoma (PDAC) has not been studied. Here, by analyzing The Cancer Genome Atlas cohort, we found that FAM84B amplification was observed in 11% of 141 PDAC patients, and FAM84B amplification was correlated with higher mRNA expression of FAM84B. FAM84B amplification and overexpression was significantly correlated with poor overall survival. Moreover, knockdown of FAM84B in PDAC cell lines suppressed cell proliferation and induced apoptosis. FAM84B knockdown also suppressed mitochondrial function and glycolysis of PDAC cells. Interestingly, knockdown of FAM84B decreased the nuclear accumulation of β-catenin, and the expression of c-Myc and lactate dehydrogenase A, but enhanced the expression of Survivin. On the contrary, FAM84B overexpression displayed reversed effects in cell proliferation, apoptosis, mitochondrial function, and glycolysis, which was blocked by the Wnt/β-catenin pathway inhibitor (XAV939). In addition, PDAC cells with lower expression of FAM84B were more sensitive to gemcitabine-induced cell proliferation inhibition both in vitro and in vivo. In conclusion, FAM84B plays an important role in aerobic glycolysis and tumorigenesis in PDAC and Wnt/β-catenin may be involved in this process.
- Published
- 2020
42. The trehalose-6-phosphate phosphatase Tps2 regulates ATG8 transcription and autophagy in Saccharomyces cerevisiae
- Author
-
Hyojeong Choi, Yongook Lee, Won-Ki Huh, and Bongkeun Kim
- Subjects
0301 basic medicine ,Autophagosome ,Saccharomyces cerevisiae Proteins ,Transcription, Genetic ,Nitrogen ,Trehalose-6-phosphate phosphatase ,ATG8 ,Saccharomyces cerevisiae ,Cellular homeostasis ,Models, Biological ,03 medical and health sciences ,chemistry.chemical_compound ,Transcription (biology) ,Catalytic Domain ,Autophagy ,Phosphorylation ,Molecular Biology ,Cell Nucleus ,030102 biochemistry & molecular biology ,biology ,Autophagy-Related Protein 8 Family ,Cell Biology ,biology.organism_classification ,Trehalose ,Cell biology ,Protein Transport ,030104 developmental biology ,chemistry ,Glucosyltransferases ,Research Paper ,Protein Binding - Abstract
Macroautophagy/autophagy is an important catabolic process for maintaining cellular homeostasis by adapting to various stress conditions. Autophagy is mediated by a double-membrane autophagosome, which sequesters a portion of cytoplasmic components for delivery to the vacuole. Several autophagy-related (ATG) genes play crucial roles in autophagosome formation. The induction of ATG genes must be tightly regulated to maintain a proper autophagic activity, but their regulatory mechanisms are still largely unknown. Here, we report that the trehalose-6-phosphate phosphatase Tps2 functions as a positive regulator of autophagy in Saccharomyces cerevisiae. Cellular trehalose levels do not affect autophagy regulation by Tps2. Loss of Tps2 leads to impaired autophagic flux and reduced ATG8 expre/ssion under nitrogen starvation. In tps2Δ cells, Ume6 is predominantly dephosphorylated and represses ATG8 transcription by binding to its promoter region. Tps2 regulates nuclear translocation and activation of Rim15 kinase, a negative regulator of Ume6, by causing the dissociation of Rim15 from the 14-3-3 proteins Bmh1/2 under nitrogen starvation, suggesting that Rim15 mediates the function of Tps2 as a positive regulator of ATG8 induction. Furthermore, Tps2 plays a crucial role in the dephosphorylation of Ser1061 and Thr1075 residues of Rim15, which is important for controlling the dissociation of Rim15 from Bmh1/2 under nitrogen starvation. Together, our results reveal the role of Tps2 as a positive regulator of autophagy and provide new insight into the regulatory mechanisms of ATG gene expression. Abbreviations: ATG: autophagy-related; ChIP: chromatin immunoprecipitation; Co-IP: co-immunoprecipitation; DAPI: 4ʹ,6-diamidino-2-phenylindole; GFP: green fluorescent protein; PKA: protein kinase A; PtdIns3K: phosphatidylinositol 3-kinase; Rim15KI: kinase-inactive Rim15; Rim15-2A: Rim15S1061A,T1075A; TEM: transmission electron microscopy; TORC1: target of rapamycin complex 1
- Published
- 2020
43. Molecular anatomy of the subcellular localization and nuclear import mechanism of herpes simplex virus 1 UL6
- Author
-
Yiwen Li, Meili Li, Yingjie Guo, Zuo Xu, Hao Peng, Mingsheng Cai, Xingmei Zou, Manjiao Lu, Weidong Gan, Yangxi Deng, Yuanfang Wang, Tao Peng, and Xiaowen Ou
- Subjects
Yellow fluorescent protein ,Aging ,Active Transport, Cell Nucleus ,Importin ,Herpesvirus 1, Human ,Viral Proteins ,FLAG-tag ,RNA interference ,Chlorocebus aethiops ,importin ,Animals ,Humans ,Cell Nucleus ,biology ,Chemistry ,Cell Biology ,Subcellular localization ,HSV-1 ,nuclear import ,Cell biology ,UL6 ,HEK293 Cells ,Ran ,Ran-GTP ,COS Cells ,biology.protein ,Nuclear transport ,Nuclear localization sequence ,Research Paper - Abstract
As an indispensable structure protein, the herpes simplex virus 1 (HSV-1) UL6 has been described to exert numerous roles in viral proliferation. However, its exact subcellular localization and subcellular transport mechanism is not well known. In the present study, by utilizing confocal fluorescent microscopy, UL6 was shown to mainly locate in the nucleus in enhanced yellow fluorescent protein or Flag tag fused expression plasmid-transfected cells or HSV-1-infected cells, whereas its predicted nuclear localization signal was nonfunctional. In addition, by exploiting dominant negative mutant and inhibitor of different nuclear import receptors, as well as co-immunoprecipitation and RNA interference assays, UL6 was established to interact with importin α1, importin α7 and transportin-1 to mediate its nuclear translocation under the help of Ran-mediated GTP hydrolysis. Accordingly, these results will advance the knowledge of UL6-mediated biological significances in HSV-1 infection cycle.
- Published
- 2020
44. SOCS1 blocks G1-S transition in hepatocellular carcinoma by reducing the stability of the CyclinD1/CDK4 complex in the nucleus
- Author
-
Haiyang Xie, Suwan Sun, Lin Zhou, Shengyong Yin, Jun Ding, Chao Qian, Wei Zhang, Kangdi Xu, and Shusen Zheng
- Subjects
Aging ,Carcinoma, Hepatocellular ,Cell Survival ,Cell ,Suppressor of cytokine signalling ,hepatocellular carcinoma (HCC) ,Mice ,Suppressor of Cytokine Signaling 1 Protein ,Cyclin-dependent kinase ,Cell Line, Tumor ,medicine ,suppressor of cytokine signalling 1 (SOCS1) ,Animals ,Cyclin D1 ,CyclinD1 ,Cell Nucleus ,biology ,Suppressor of cytokine signaling 1 ,Chemistry ,Cell growth ,business.industry ,Cell Cycle ,Liver Neoplasms ,Cancer ,Cyclin-Dependent Kinase 4 ,G1/S transition ,Cell Biology ,Cell cycle ,medicine.disease ,medicine.anatomical_structure ,cell proliferation ,Hepatocellular carcinoma ,biology.protein ,Cancer research ,Hepatocytes ,Heterografts ,business ,Research Paper - Abstract
Background: Inhibitors of the CDK family of proteins have been approved for the treatment of a variety of tumours; however, the use of this type of drug alone is often ineffective and elicits adverse side effects. The development of new drugs administered in combination with CDK inhibitors is expected to improve the therapeutic effect. Methods: We identified the function of suppressor of cytokine signalling 1 (SOCS1) in hepatocellular carcinoma (HCC) cell models and the xenograft mouse model. The molecular mechanism of SOCS1 regulating cell proliferation was proved through transcriptome sequencing, western blot and Co-IP. Findings: When SOCS1 expression was artificially upregulated, HCC cell lines were arrested at the G1-S transition in the cell cycle. Interestingly, during this process, total CyclinD1 protein increased, but the effective proportion decreased. We found that the deficiency of CyclinD1 in the nucleus is probably due to the decrease in the stability of nuclear CyclinD1 caused by the ubiquitin-based degradation of P21 and P27, thus inhibiting the progression of the cell cycle to S phase. After P21 expression was increased, the levels of the component that inactivates CyclinD1 decreased as expected. It showed that P21 has a partial promoting effect on cancer. Interpretation: As a tumour inhibitory factor in HCC, SOCS1 is a good indicator of prognosis, tumour size and long-term survival after resection. SOCS1 is expected to become a drug target in combined with CDK family inhibitors. Funding Statement: Innovative Research Groups of National Natural Science Foundation of China (No. 81721091), Major program of National Natural Science Foundation of China (No.91542205), National S&T Major Project (No. 2017ZX10203205), Zhejiang International Science and Technology Cooperation Project (No.2016C04003), National Natural Science Foundation of China (No. 81773070). Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The study protocol was approved by the ethics committee of the First Affiliated Hospital of Zhejiang University. All experiments containing animals according to protocols approved by Animal Care and Use Committee of Zhejiang University.
- Published
- 2020
45. The nuclear localization signal-mediated nuclear targeting of herpes simplex virus 1 early protein UL2 is important for efficient viral production
- Author
-
Xingmei Zou, Yilong Bao, Xiaowen Ou, Tong Li, Meili Li, Yangxi Deng, Mingsheng Cai, Yingjie Guo, Tao Peng, Yiwen Li, Yuanfang Wang, Zuo Xu, Shaoxuan Shi, Delong Liu, and Si Jiang
- Subjects
Gene Expression Regulation, Viral ,Aging ,viruses ,DNA, Recombinant ,Herpesvirus 1, Human ,Viral Plaque Assay ,Biology ,medicine.disease_cause ,Recombinant virus ,Virus Replication ,Viral Proteins ,Chlorocebus aethiops ,medicine ,NLS ,Animals ,Humans ,HSV-1 UL2 ,Nuclear export signal ,Vero Cells ,Cell Nucleus ,Mutation ,DNA replication ,Cell Biology ,nuclear localization signal ,Cell biology ,Protein Transport ,Herpes simplex virus ,HEK293 Cells ,COS Cells ,DNA, Viral ,recombinant virus ,Homologous recombination ,Nuclear localization sequence ,Gene Deletion ,Research Paper - Abstract
Herpes simplex virus 1 (HSV-1) is a representative alphaherpesvirus that can provoke a series of severe diseases to human being, but its exact pathogenesis is not perfectly understood. UL2, a uracil-DNA glycosylase involved in the process of HSV-1 DNA replication, has been shown to be predominantly targeted to the nuclei in our previous study, yet little is established regarding the subcellular localization signal or its related function of UL2 during HSV-1 propagation. Here, by creating a number of UL2 variants merged with enhanced yellow fluorescent protein, an authentic nuclear localization signal (NLS) of UL2 was, for the first time, identified and profiled to amino acids (aa) 1 to 17 (MKRACSRSPSPRRRPSS), and 12RRR14 was indispensable for its nuclear accumulation. Besides, the predicted nuclear export signal (aa 225 to 240) of UL2 was determined to be nonfunctional. Based on the HSV-1 bacterial artificial chromosome and homologous recombination technique, three recombinant viruses with mutations of the identified NLS, deletion and revertant of UL2 were constructed to assess the effect of UL2 nuclear targeting on HSV-1 replication. Compared to the wild type HSV-1, UL2 deletion remarkably restrained viral production, and mutation of NLS targeting UL2 to cytoplasm (pan-cellular distribution) in recombinant virus-infected cells showed a certain degree of deficiency in HSV-1 proliferation. Moreover, recombinant virus with UL2 deletion exhibited serious damages of viral DNA synthesis and mRNA expression, and these processes were partially disrupted in the recombinant virus with UL2 NLS mutation. Collectively, we had established a functional NLS in UL2 and showed that the NLS-mediated nuclear translocation of UL2 was important for efficient production of HSV-1. These data were of significance for further clarifying the biological function of UL2 during HSV-1 infection.
- Published
- 2020
46. Development of CAPER peptides for the treatment of triple negative breast cancer
- Author
-
Sundee Dees, Isaac Hutchinson, Laura Pontiggia, Rachel Trimble, Shannon D Chilewski, Isabelle Mercier, Jean-Francois Jasmin, and Devyani Bhosale
- Subjects
Cyclin-Dependent Kinase Inhibitor p21 ,0301 basic medicine ,Carcinogenesis ,Cell Survival ,Proto-Oncogene Proteins c-jun ,DNA repair ,Estrogen receptor ,Apoptosis ,Cell Count ,Triple Negative Breast Neoplasms ,CAPER ,Biology ,RNA binding protein-39 (Rbm39) ,Histones ,03 medical and health sciences ,0302 clinical medicine ,Breast cancer ,Cell Line, Tumor ,Coactivator ,Progesterone receptor ,medicine ,Humans ,Cyclin D1 ,Triple negative breast cancer ,Amino Acid Sequence ,Phosphorylation ,Receptor ,Molecular Biology ,Triple-negative breast cancer ,Cell Nucleus ,peptide therapeutics ,Cell Cycle ,c-Jun ,Cell Biology ,medicine.disease ,Recombinant Proteins ,Neoplasm Proteins ,030104 developmental biology ,Proto-Oncogene Proteins c-bcl-2 ,030220 oncology & carcinogenesis ,Cancer research ,hepatocellular carcinoma-1.4 (HCC1.4) ,Peptides ,Protein Binding ,Research Paper ,Developmental Biology - Abstract
Triple negative breast cancer (TNBC) is a heterogeneous disease, which lacks expression of the estrogen receptor (ER), progesterone receptor (PR) and the human epidermal growth factor 2 receptor (HER2). This subtype of breast cancer has the poorest prognosis with limited therapies currently available, and hence additional options are needed. CAPER is a coactivator of the activator protein-1 (AP-1) (interacting specifically with the c-Jun component) and the ER and is known to be involved in human breast cancer pathogenesis. Recent published data have demonstrated a role for CAPER in TNBC and, as such, disrupting the function of CAPER with c-Jun could be a novel approach to treat TNBC patients. The data presented here shows the development and in vitro testing of CAPER-derived peptides that inhibit the coactivator activity of CAPER with c-Jun. These CAPER peptides result in a decrease in cell number and an increase in apoptosis in two TNBC cell lines, MDA-MB-231 and BT-549, while having no effect on the non-tumorigenic cell line MCF 10A. Additionally, two modes of action were demonstrated which appear to be cell line dependent: 1) a modulation of phosphorylated c-Jun leading to a decrease in Bcl-2 in MDA-MB-231 cells and a decrease in p21 in BT-549 cells and 2) a decrease in DNA repair proteins, leading to impaired DNA repair function in MDA-MB-231 cells. The data presented here supports further development of CAPER-derived peptides for the treatment of TNBC.
- Published
- 2020
47. Upon microbial challenge, human neutrophils undergo rapid changes in nuclear architecture and chromatin folding to orchestrate an immediate inflammatory gene program
- Author
-
Cornelis Murre, Victor Nizet, Yi-Na Zhu, Hanbin Lu, Zhaoren He, Takeshi Isoda, Matthew Denholtz, and Simon Döhrmann
- Subjects
Transcriptional Activation ,Neutrophils ,Biology ,Proinflammatory cytokine ,Histones ,03 medical and health sciences ,0302 clinical medicine ,Gene expression ,Escherichia coli ,Genetics ,CEBPB ,Humans ,Compartment (development) ,Enhancer ,Cells, Cultured ,Escherichia coli Infections ,030304 developmental biology ,Cell Nucleus ,0303 health sciences ,Cohesin ,FOSL2 ,Chromatin ,Cell biology ,030220 oncology & carcinogenesis ,Research Paper ,Developmental Biology - Abstract
Differentiating neutrophils undergo large-scale changes in nuclear morphology. How such alterations in structure are established and modulated upon exposure to microbial agents is largely unknown. Here, we found that prior to encounter with bacteria, an armamentarium of inflammatory genes was positioned in a transcriptionally passive environment suppressing premature transcriptional activation. Upon microbial exposure, however, human neutrophils rapidly (
- Published
- 2020
48. Single-cell RNA-sequencing analysis identifies host long noncoding RNA MAMDC2-AS1 as a co-factor for HSV-1 nuclear transport
- Author
-
Ji Xiao, Yun Wang, Wang Yifei, Fujun Jin, Xiaowei Song, Lianzhou Huang, Weisheng Luo, Yiliang Wang, Zhaoyang Wang, Yuan Wang, Feng Li, and Shurong Qin
- Subjects
Hsp90α ,viruses ,Active Transport, Cell Nucleus ,RNA-binding protein ,Herpesvirus 1, Human ,MAMDC2-AS1 ,nuclear transport ,Biology ,Applied Microbiology and Biotechnology ,Cell Line ,03 medical and health sciences ,Transcription (biology) ,Gene expression ,Humans ,Gene silencing ,HSP90 Heat-Shock Proteins ,RNA-Seq ,single-cell RNA-sequencing ,Genes, Immediate-Early ,Molecular Biology ,Transcription factor ,Gene ,YY1 Transcription Factor ,Ecology, Evolution, Behavior and Systematics ,030304 developmental biology ,Cell Nucleus ,0303 health sciences ,RNA-Binding Proteins ,RNA ,Herpes Simplex Virus Protein Vmw65 ,Cell Biology ,HSV-1 ,tegument protein ,Cell biology ,RNA, Long Noncoding ,Ectopic expression ,Single-Cell Analysis ,Research Paper ,Developmental Biology - Abstract
Herpes simplex virus (HSV) type 1 (HSV-1) infection exhibited high heterogeneity at individual cells level, including the different gene expression patterns and varying amounts of progeny virus. However, the underlying mechanism of such variability remains obscure. The importance of host long noncoding RNAs (lncRNAs) in virus infection had been recognized, while the contribution of lncRNAs to the heterogeneous infection remains unknown. Herein, a prior single-cell RNA sequencing data using HSV-1 reporter strain expressing ICP4-YFP was re-analyzed to obtain the differentially expressed lncRNA between the successfully initiated viral gene expression (ICP4-YFP+) cells and the aborted infection cells (ICP4-YFP-). The ICP4-YFP+ population show a higher abundance of MAMDC2 antisense 1 (MAMDC2-AS1) lncRNA than ICP4-YFP- population. MAMDC2-AS1 silencing reduces the expression of HSV-1 immediate early (IE) genes and limit HSV-1 infection in human host cells. Consistently, ectopic expression of MAMDC2-AS1 enhances HSV-1 IE genes transcription and facilitates the formation of HSV-1-induced plaques. Mechanically, both RNA-pull down and RNA immunoprecipitation assays show that MAMDC2-AS1 interacts with the RNA binding protein heat shock protein 90α (Hsp90α), a molecular chaperone involving in the nuclear import of HSV-1. The MAMDC2-AS1-Hsp90α interaction facilitates the nuclear transport of viral tegument protein VP16, the core factor initiating the expression of HSV-1 IE genes. The transcription factor YY1 mediates the induction of MAMDC2-AS1 upon HSV-1 infection. Our study elucidates the contribution of lncRNA to HSV-1 infection susceptibility in human cells and the role of Hsp90α RNA binding activity in HSV-1 infection.
- Published
- 2020
49. Unexplored Cdc42 functions at the budding yeast nucleus suggested by subcellular localization
- Author
-
David G. Drubin and Michelle S. Lu
- Subjects
rho GTP-Binding Proteins ,Saccharomyces cerevisiae Proteins ,nucleopodia ,1.1 Normal biological development and functioning ,macromolecular substances ,GTPase ,Vacuole ,CDC42 ,Saccharomyces cerevisiae ,Biology ,Biochemistry ,nucleoporins ,Underpinning research ,medicine ,Cell Nucleus ,vacuole ,Cell Polarity ,Cell Biology ,Pharmacology and Pharmaceutical Sciences ,Cell cycle ,Subcellular localization ,Cell biology ,Nucleus-vacuole junction ,medicine.anatomical_structure ,Saccharomycetales ,nucleus-vacuole junction ,Nucleoporin ,Generic health relevance ,Biochemistry and Cell Biology ,biological phenomena, cell phenomena, and immunity ,Nucleus ,Cell Division ,Research Paper - Abstract
In budding yeast, the Rho-family GTPase Cdc42 has several functions that depend on its subcellular localization and the cell cycle stage. During bud formation, Cdc42 localizes to the plasma membrane at the bud tip and bud neck where it carries out functions in actin polymerization, spindle positioning, and exocytosis to ensure proper polarity development. Recent live-cell imaging analysis revealed a novel localization of Cdc42 to a discrete intracellular focus associated with the vacuole and nuclear envelope. The discovery of this novel Cdc42 localization led to the identification of a new function in ESCRT-mediated nuclear envelope sealing. However, other aspects of this intracellular localization and its functional implications were not explored. Here, we further characterize the Cdc42 focus and present several novel observations that suggest possible additional Cdc42 functions at the nucleus, including nucleus-vacuole junction formation, nuclear envelope tethering, nuclear migration, and nucleopodia formation.
- Published
- 2022
50. Nucleocytoplasmic shuttling of the GPN-loop GTPase Gpn3 is regulated by serum and cell density in MCF-12A mammary cells.
- Author
-
Peña-Gómez SG, Cristóbal-Mondragón GR, Vega-Palomo CR, Mora-García M, Félix-Pérez T, Rebolloso-Gómez Y, Calera MR, and Sánchez-Olea R
- Subjects
- Active Transport, Cell Nucleus, Proteasome Endopeptidase Complex metabolism, Cell Count, GTP Phosphohydrolases metabolism, Cell Nucleus metabolism
- Abstract
The best-known function of the essential GPN-loop GTPase Gpn3 is to contribute to RNA polymerase II assembly, a prerequisite for its nuclear targeting. Although this process occurs in the cytoplasm, we have previously shown that Gpn3 enters the cell nucleus before being polyubiquitinated. Here, we show that inhibiting Crm1-mediated nuclear export with leptomycin B, or the proteasome with MG132, caused the nuclear accumulation of recombinant and endogenous Gpn3 in MCF-12A cells. When added simultaneously, leptomycin B and MG132 had an additive effect. Analysis of Gpn3 primary sequence revealed the presence of at least five nuclear export sequence (NES) motifs, with some having a higher exposure to the solvent in the GTP-bound than GDP-bound state in a Gpn3 structural model. Inactivation of any of these NESes led to some degree of Gpn3 nuclear accumulation, although mutating NES1 or NES3 had the more robust effect. MCF-12A cells expressing exclusively a NES-deficient version of Gpn3R-Flag proliferated slower than cells expressing Gpn3R-Flag wt, indicating that nuclear export is important for Gpn3 function. Next, we searched for physiological conditions regulating Gpn3 nucleocytoplasmic shuttling. Interestingly, whereas Gpn3R-Flag was both nuclear and cytoplasmic in low-density growing MCF-12A cells, it was exclusively cytoplasmic in high-density areas. Furthermore, Gpn3R-Flag was cytoplasmic, mostly perinuclear, in sparse but starved MCF-12A cells, and serum-stimulation caused a rapid, although transient, Gpn3R-Flag nuclear accumulation. We conclude that Gpn3 nucleocytoplasmic shuttling is regulated by cell density and growth factors, and propose that Gpn3 has an unknown nuclear function positively linked to cell growth and/or proliferation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)
- Published
- 2024
- Full Text
- View/download PDF
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.