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1.
Nat Commun ; 14(1): 3244, 2023 06 05.
Article in English | MEDLINE | ID: covidwho-20239143

ABSTRACT

Variations of cell-type proportions within tissues could be informative of biological aging and disease risk. Single-cell RNA-sequencing offers the opportunity to detect such differential abundance patterns, yet this task can be statistically challenging due to the noise in single-cell data, inter-sample variability and because such patterns are often of small effect size. Here we present a differential abundance testing paradigm called ELVAR that uses cell attribute aware clustering when inferring differentially enriched communities within the single-cell manifold. Using simulated and real single-cell and single-nucleus RNA-Seq datasets, we benchmark ELVAR against an analogous algorithm that uses Louvain for clustering, as well as local neighborhood-based methods, demonstrating that ELVAR improves the sensitivity to detect cell-type composition shifts in relation to aging, precancerous states and Covid-19 phenotypes. In effect, leveraging cell attribute information when inferring cell communities can denoise single-cell data, avoid the need for batch correction and help retrieve more robust cell states for subsequent differential abundance testing. ELVAR is available as an open-source R-package.


Subject(s)
COVID-19 , Single-Cell Gene Expression Analysis , Humans , Single-Cell Analysis/methods , RNA-Seq/methods , Algorithms , Cluster Analysis , Sequence Analysis, RNA/methods , Gene Expression Profiling/methods
2.
Biochim Biophys Acta Mol Basis Dis ; 1869(5): 166707, 2023 06.
Article in English | MEDLINE | ID: covidwho-2269405

ABSTRACT

INTRODUCTION: The COVID-19 pandemic provide the opportunities to explore the numerous similarities in clinical symptoms with Kawasaki disease (KD), including severe vasculitis. Despite this, the underlying mechanisms of vascular injury in both KD and COVID-19 remain elusive. To identify these mechanisms, this study employs single-cell RNA sequencing to explore the molecular mechanisms of immune responses in vasculitis, and validate the results through in vitro experiments. METHOD: The single-cell RNA sequencing (scRNA-seq) analysis of peripheral blood mononuclear cells (PBMCs) was carried out to investigate the molecular mechanisms of immune responses in vasculitis in KD and COVID-19. The analysis was performed on PBMCs from six children diagnosed with complete KD, three age-matched KD healthy controls (KHC), six COVID-19 patients (COV), three influenza patients (FLU), and four healthy controls (CHC). The results from the scRNA-seq analysis were validated through flow cytometry and immunofluorescence experiments on additional human samples. Subsequently, monocyte adhesion assays, immunofluorescence, and quantitative polymerase chain reaction (qPCR) were used to analyze the damages to endothelial cells post-interaction with monocytes in HUVEC and THP1 cultures. RESULTS: The scRNA-seq analysis revealed the potential cellular types involved and the alterations in genetic transcriptions in the inflammatory responses. The findings indicated that while the immune cell compositions had been altered in KD and COV patients, and the ratio of CD14+ monocytes were both elevated in KD and COV. While the CD14+ monocytes share a large scale of same differentiated expressed geens between KD and COV. The differential activation of CD14 and CD16 monocytes was found to respond to both endothelial and epithelial dysfunctions. Furthermore, SELL+/CCR1+/XAF1+ CD14 monocytes were seen to enhance the adhesion and damage to endothelial cells. The results also showed that different types of B cells were involved in both KD and COV, while only the activation of T cells was recorded in KD. CONCLUSION: In conclusion, our study demonstrated the role of the innate immune response in the regulation of endothelial dysfunction in both KD and COVID-19. Additionally, our findings indicate that the adaptive immunity activation differs between KD and COVID-19. Our results demonstrate that monocytes in COVID-19 exhibit adhesion to both endothelial cells and alveolar epithelial cells, thus providing insight into the mechanisms and shared phenotypes between KD and COVID-19.


Subject(s)
COVID-19 , Mucocutaneous Lymph Node Syndrome , Vasculitis , Child , Humans , Monocytes/metabolism , Mucocutaneous Lymph Node Syndrome/genetics , Mucocutaneous Lymph Node Syndrome/metabolism , Leukocytes, Mononuclear/metabolism , Endothelial Cells/metabolism , Pandemics , RNA-Seq , Lipopolysaccharide Receptors/metabolism , COVID-19/metabolism , Vasculitis/genetics , Vasculitis/metabolism , Receptors, CCR1
3.
Cells ; 12(3)2023 01 21.
Article in English | MEDLINE | ID: covidwho-2289161

ABSTRACT

Genes with similar expression patterns in a set of diverse samples may be considered coexpressed. Human Gene Coexpression Analysis 2.0 (HGCA2.0) is a webtool which studies the global coexpression landscape of human genes. The website is based on the hierarchical clustering of 55,431 Homo sapiens genes based on a large-scale coexpression analysis of 3500 GTEx bulk RNA-Seq samples of healthy individuals, which were selected as the best representative samples of each tissue type. HGCA2.0 presents subclades of coexpressed genes to a gene of interest, and performs various built-in gene term enrichment analyses on the coexpressed genes, including gene ontologies, biological pathways, protein families, and diseases, while also being unique in revealing enriched transcription factors driving coexpression. HGCA2.0 has been successful in identifying not only genes with ubiquitous expression patterns, but also tissue-specific genes. Benchmarking showed that HGCA2.0 belongs to the top performing coexpression webtools, as shown by STRING analysis. HGCA2.0 creates working hypotheses for the discovery of gene partners or common biological processes that can be experimentally validated. It offers a simple and intuitive website design and user interface, as well as an API endpoint.


Subject(s)
Gene Expression Profiling , Gene Regulatory Networks , Genes , Humans , RNA-Seq , Transcription Factors , Genes/genetics , Genes/physiology
4.
Cytokine ; 166: 156187, 2023 06.
Article in English | MEDLINE | ID: covidwho-2279243

ABSTRACT

COVID-19 is associated with dysregulation of several genes and signaling pathways. Based on the importance of expression profiling in identification of the pathogenesis of COVID-19 and proposing novel therapies for this disorder, we have employed an in silico approach to find differentially expressed genes between COVID-19 patients and healthy controls and their relevance with cellular functions and signaling pathways. We obtained 630 DEmRNAs, including 486 down-regulated DEGs (such as CCL3 and RSAD2) and 144 up-regulated DEGs (such as RHO and IQCA1L), and 15 DElncRNAs, including 9 down-regulated DElncRNAs (such as PELATON and LINC01506) and 6 up-regulated DElncRNAs (such as AJUBA-DT and FALEC). The PPI network of DEGs showed the presence of a number immune-related genes such as those coding for HLA molecules and interferon regulatory factors. Taken together, these results highlight the importance of immune-related genes and pathways in the pathogenesis of COVID-19 and suggest novel targets for treatment of this disorder.


Subject(s)
COVID-19 , Gene Expression Profiling , Humans , Gene Expression Profiling/methods , Systems Biology , SARS-CoV-2/genetics , Computational Biology/methods , COVID-19/genetics , RNA-Seq , LIM Domain Proteins
5.
Mil Med Res ; 9(1): 68, 2022 12 02.
Article in English | MEDLINE | ID: covidwho-2196508

ABSTRACT

The application of single-cell RNA sequencing (scRNA-seq) in biomedical research has advanced our understanding of the pathogenesis of disease and provided valuable insights into new diagnostic and therapeutic strategies. With the expansion of capacity for high-throughput scRNA-seq, including clinical samples, the analysis of these huge volumes of data has become a daunting prospect for researchers entering this field. Here, we review the workflow for typical scRNA-seq data analysis, covering raw data processing and quality control, basic data analysis applicable for almost all scRNA-seq data sets, and advanced data analysis that should be tailored to specific scientific questions. While summarizing the current methods for each analysis step, we also provide an online repository of software and wrapped-up scripts to support the implementation. Recommendations and caveats are pointed out for some specific analysis tasks and approaches. We hope this resource will be helpful to researchers engaging with scRNA-seq, in particular for emerging clinical applications.


Subject(s)
Biomedical Research , Data Analysis , Humans , RNA-Seq
6.
FEBS J ; 290(12): 3128-3144, 2023 06.
Article in English | MEDLINE | ID: covidwho-2192593

ABSTRACT

Viral infections can modulate the widespread alternations of cellular splicing, favouring viral replication within the host cells by overcoming host immune responses. However, how SARS-CoV-2 induces host cell differential splicing and affects the landscape of transcript alternation in severe COVID-19 infection remains elusive. Understanding the differential splicing and transcript alternations in severe COVID-19 infection may improve our molecular insights into the SARS-CoV-2 pathogenesis. In this study, we analysed the publicly available blood and lung transcriptome data of severe COVID-19 patients, blood transcriptome data of recovered COVID-19 patients at 12-, 16- and 24-week postinfection and healthy controls. We identified a significant transcript isoform switching in the individual blood and lung RNA-seq data of severe COVID-19-infected patients and 25 common genes that alter their transcript isoform in both blood and lung samples. Altered transcripts show significant loss of the open reading frame, functional domains and switch from coding to noncoding transcript, impacting normal cellular functions. Furthermore, we identified the expression of several novel recurrent chimeric transcripts in the blood samples from severe COVID-19 patients. Moreover, the analysis of the isoform switching into blood samples from recovered COVID-19 patients highlights that there is no significant isoform switching in 16- and 24-week postinfection, and the levels of expressed chimeric transcripts are reduced. This finding emphasizes that SARS-CoV-2 severe infection induces widespread splicing in the host cells, which could help the virus alter the host immune responses and facilitate the viral replication within the host and the efficient translation of viral proteins.


Subject(s)
COVID-19 , Humans , SARS-CoV-2/genetics , Lung/metabolism , Transcriptome , RNA-Seq
7.
Int J Mol Sci ; 23(24)2022 Dec 10.
Article in English | MEDLINE | ID: covidwho-2155138

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly contagious and pathogenic coronavirus that emerged in late 2019 and caused a pandemic of respiratory illness termed as coronavirus disease 2019 (COVID-19). Cancer patients are more susceptible to SARS-CoV-2 infection. The treatment of cancer patients infected with SARS-CoV-2 is more complicated, and the patients are at risk of poor prognosis compared to other populations. Patients infected with SARS-CoV-2 are prone to rapid development of acute respiratory distress syndrome (ARDS) of which pulmonary fibrosis (PF) is considered a sequelae. Both ARDS and PF are factors that contribute to poor prognosis in COVID-19 patients. However, the molecular mechanisms among COVID-19, ARDS and PF in COVID-19 patients with cancer are not well-understood. In this study, the common differentially expressed genes (DEGs) between COVID-19 patients with and without cancer were identified. Based on the common DEGs, a series of analyses were performed, including Gene Ontology (GO) and pathway analysis, protein-protein interaction (PPI) network construction and hub gene extraction, transcription factor (TF)-DEG regulatory network construction, TF-DEG-miRNA coregulatory network construction and drug molecule identification. The candidate drug molecules (e.g., Tamibarotene CTD 00002527) obtained by this study might be helpful for effective therapeutic targets in COVID-19 patients with cancer. In addition, the common DEGs among ARDS, PF and COVID-19 patients with and without cancer are TNFSF10 and IFITM2. These two genes may serve as potential therapeutic targets in the treatment of COVID-19 patients with cancer. Changes in the expression levels of TNFSF10 and IFITM2 in CD14+/CD16+ monocytes may affect the immune response of COVID-19 patients. Specifically, changes in the expression level of TNFSF10 in monocytes can be considered as an immune signature in COVID-19 patients with hematologic cancer. Targeting N6-methyladenosine (m6A) pathways (e.g., METTL3/SERPINA1 axis) to restrict SARS-CoV-2 reproduction has therapeutic potential for COVID-19 patients.


Subject(s)
COVID-19 , Neoplasms , Pulmonary Fibrosis , Respiratory Distress Syndrome , Humans , COVID-19/complications , COVID-19/genetics , Lung/pathology , Membrane Proteins/metabolism , Methyltransferases/metabolism , Neoplasms/complications , Neoplasms/genetics , Pulmonary Fibrosis/pathology , Pulmonary Fibrosis/virology , Respiratory Distress Syndrome/pathology , Respiratory Distress Syndrome/virology , RNA-Seq , SARS-CoV-2 , Single-Cell Gene Expression Analysis , Transcription Factors/metabolism
8.
PLoS One ; 17(10): e0276460, 2022.
Article in English | MEDLINE | ID: covidwho-2089429

ABSTRACT

Excessive neutrophil infiltration and dysfunction contribute to the progression and severity of hyper-inflammatory syndrome, such as in severe COVID19. In the current study, we re-analysed published scRNA-seq datasets of mouse and human neutrophils to classify and compare the transcriptional regulatory networks underlying neutrophil differentiation and inflammatory responses. Distinct sets of TF modules regulate neutrophil maturation, function, and inflammatory responses under the steady state and inflammatory conditions. In COVID19 patients, neutrophil activation was associated with the selective activation of inflammation-specific TF modules. SARS-CoV-2 RNA-positive neutrophils showed a higher expression of type I interferon response TF IRF7. Furthermore, IRF7 expression was abundant in neutrophils from severe patients in progression stage. Neutrophil-mediated inflammatory responses positively correlate with the expressional level of IRF7. Based on these results, we suggest that differential activation of activation-related TFs, such as IRF7 mediate neutrophil inflammatory responses during inflammation.


Subject(s)
COVID-19 , Neutrophils , Humans , COVID-19/genetics , COVID-19/metabolism , Inflammation/genetics , Interferon Type I/metabolism , Neutrophil Activation/genetics , Neutrophil Activation/physiology , Neutrophils/metabolism , RNA, Viral , RNA-Seq , SARS-CoV-2 , Single-Cell Analysis
9.
Brief Bioinform ; 23(5)2022 09 20.
Article in English | MEDLINE | ID: covidwho-2017730

ABSTRACT

We present a novel self-supervised Contrastive LEArning framework for single-cell ribonucleic acid (RNA)-sequencing (CLEAR) data representation and the downstream analysis. Compared with current methods, CLEAR overcomes the heterogeneity of the experimental data with a specifically designed representation learning task and thus can handle batch effects and dropout events simultaneously. It achieves superior performance on a broad range of fundamental tasks, including clustering, visualization, dropout correction, batch effect removal, and pseudo-time inference. The proposed method successfully identifies and illustrates inflammatory-related mechanisms in a COVID-19 disease study with 43 695 single cells from peripheral blood mononuclear cells.


Subject(s)
COVID-19 , RNA , COVID-19/genetics , Cluster Analysis , Data Analysis , Humans , Leukocytes, Mononuclear , RNA-Seq , Sequence Analysis, RNA/methods
10.
Proc Natl Acad Sci U S A ; 119(36): e2120680119, 2022 09 06.
Article in English | MEDLINE | ID: covidwho-2001001

ABSTRACT

The systemic immune response to viral infection is shaped by master transcription factors, such as NF-κB, STAT1, or PU.1. Although long noncoding RNAs (lncRNAs) have been suggested as important regulators of transcription factor activity, their contributions to the systemic immunopathologies observed during SARS-CoV-2 infection have remained unknown. Here, we employed a targeted single-cell RNA sequencing approach to reveal lncRNAs differentially expressed in blood leukocytes during severe COVID-19. Our results uncover the lncRNA PIRAT (PU.1-induced regulator of alarmin transcription) as a major PU.1 feedback-regulator in monocytes, governing the production of the alarmins S100A8/A9, key drivers of COVID-19 pathogenesis. Knockout and transgene expression, combined with chromatin-occupancy profiling, characterized PIRAT as a nuclear decoy RNA, keeping PU.1 from binding to alarmin promoters and promoting its binding to pseudogenes in naïve monocytes. NF-κB-dependent PIRAT down-regulation during COVID-19 consequently releases a transcriptional brake, fueling alarmin production. Alarmin expression is additionally enhanced by the up-regulation of the lncRNA LUCAT1, which promotes NF-κB-dependent gene expression at the expense of targets of the JAK-STAT pathway. Our results suggest a major role of nuclear noncoding RNA networks in systemic antiviral responses to SARS-CoV-2 in humans.


Subject(s)
COVID-19 , Gene Expression Regulation , Monocytes , RNA, Long Noncoding , SARS-CoV-2 , Alarmins/genetics , COVID-19/genetics , COVID-19/immunology , Humans , Janus Kinases/genetics , Monocytes/immunology , NF-kappa B/genetics , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , RNA-Seq , SARS-CoV-2/immunology , STAT Transcription Factors/genetics , Signal Transduction/genetics , Single-Cell Analysis
11.
Nature ; 609(7928): 754-760, 2022 09.
Article in English | MEDLINE | ID: covidwho-1984401

ABSTRACT

Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge1-5. Here we conducted a genome-wide association study (GWAS) involving 2,393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3,289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target.


Subject(s)
COVID-19 , GTPase-Activating Proteins , Genome-Wide Association Study , Guanine Nucleotide Exchange Factors , Host Microbial Interactions , SARS-CoV-2 , Alleles , Animals , COVID-19/complications , COVID-19/genetics , COVID-19/immunology , COVID-19/physiopathology , Disease Models, Animal , GTPase-Activating Proteins/antagonists & inhibitors , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , Genetic Predisposition to Disease , Guanine Nucleotide Exchange Factors/antagonists & inhibitors , Guanine Nucleotide Exchange Factors/genetics , Guanine Nucleotide Exchange Factors/metabolism , Host Microbial Interactions/genetics , Host Microbial Interactions/immunology , Humans , Interferon Type I/genetics , Interferon Type I/immunology , Japan , Lung/pathology , Macrophages , Mesocricetus , Middle Aged , Pneumonia/complications , Pyrazoles/pharmacology , RNA-Seq , SARS-CoV-2/pathogenicity , Viral Load , Weight Loss
12.
Proc Natl Acad Sci U S A ; 119(33): e2203437119, 2022 08 16.
Article in English | MEDLINE | ID: covidwho-1960624

ABSTRACT

The mortality of coronavirus disease 2019 (COVID-19) is strongly correlated with pulmonary vascular pathology accompanied by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection-triggered immune dysregulation and aberrant activation of platelets. We combined histological analyses using field emission scanning electron microscopy with energy-dispersive X-ray spectroscopy analyses of the lungs from autopsy samples and single-cell RNA sequencing of peripheral blood mononuclear cells to investigate the pathogenesis of vasculitis and immunothrombosis in COVID-19. We found that SARS-CoV-2 accumulated in the pulmonary vessels, causing exudative vasculitis accompanied by the emergence of thrombospondin-1-expressing noncanonical monocytes and the formation of myosin light chain 9 (Myl9)-containing microthrombi in the lung of COVID-19 patients with fatal disease. The amount of plasma Myl9 in COVID-19 was correlated with the clinical severity, and measuring plasma Myl9 together with other markers allowed us to predict the severity of the disease more accurately. This study provides detailed insight into the pathogenesis of vasculitis and immunothrombosis, which may lead to optimal medical treatment for COVID-19.


Subject(s)
COVID-19 , Lung , Myosin Light Chains , SARS-CoV-2 , Severity of Illness Index , Thromboinflammation , Vasculitis , COVID-19/blood , COVID-19/complications , COVID-19/pathology , Humans , Leukocytes, Mononuclear , Lung/blood supply , Lung/metabolism , Lung/pathology , Lung/virology , Myosin Light Chains/blood , RNA-Seq , SARS-CoV-2/isolation & purification , Single-Cell Analysis , Spectrometry, X-Ray Emission , Thromboinflammation/pathology , Thromboinflammation/virology , Vasculitis/pathology , Vasculitis/virology
13.
Comput Biol Med ; 147: 105684, 2022 08.
Article in English | MEDLINE | ID: covidwho-1930823

ABSTRACT

BACKGROUND: The world has been battling the continuous COVID-19 pandemic spread by the SARS-CoV-2 virus for last two years. The issue of viral disease prediction is constantly a matter of interest in virology and the study of disease transmission over the long years. OBJECTIVE: In this study, we aimed to implement genome association studies using RNA-Seq of COVID-19 and reveal highly expressed gene biomarkers and prediction based on the machine learning model of COVID-19 analysis to combat this pandemic. METHOD: We collected RNA-Seq gene count data for both healthy (Control) and non-healthy (Treated) COVID-19 cases. In this experiment, a sequence of bioinformatics strategies and statistical techniques, such as fold-change and adjusted p-value, were processed to identify differentially expressed genes (DEGs). We filtered biomarker sets of high DEGs, moderate DEGs, and low DEGs using DESeq2, Limma Trend, and Limma Voom methods based on intersection and union operations and applied machine learning techniques to predict COVID-19. RESULT: Through experimental analysis, 67 potential biomarkers were extracted, comprising 49 up-regulated and 18 down-regulated genes, using statistical techniques and a set-theory consensus strategy. We trained the machine learning models on 12 different biomarker sets and found that the SVM model performed better than the other classifiers with 99.07% classification accuracy for moderate DEGs. CONCLUSION: Our study revealed that identified differentially expressed genes of the moderate DEGs biomarker set, |log2FC| ≥ 2 with adjusted p-value < 0.05, work significantly as input features to implement a machine learning model using a kernel-based SVM technique to predict COVID-19.


Subject(s)
COVID-19 , Biomarkers , COVID-19/diagnosis , COVID-19/genetics , Humans , Pandemics , RNA-Seq , SARS-CoV-2/genetics
14.
Front Immunol ; 13: 798712, 2022.
Article in English | MEDLINE | ID: covidwho-1779939

ABSTRACT

The immune system is a complex and sophisticated biological system, spanning multiple levels of complexity, from the molecular level to that of tissue. Our current understanding of its function and complexity, of the heterogeneity of leukocytes, is a result of decades of concentrated efforts to delineate cellular markers using conventional methods of antibody screening and antigen identification. In mammalian models, this led to in-depth understanding of individual leukocyte subsets, their phenotypes, and their roles in health and disease. The field was further propelled forward by the development of single-cell (sc) RNA-seq technologies, offering an even broader and more integrated view of how cells work together to generate a particular response. Consequently, the adoption of scRNA-seq revealed the unexpected plasticity and heterogeneity of leukocyte populations and shifted several long-standing paradigms of immunology. This review article highlights the unprecedented opportunities offered by scRNA-seq technology to unveil the individual contributions of leukocyte subsets and their crosstalk in generating the overall immune responses in bony fishes. Single-cell transcriptomics allow identifying unseen relationships, and formulating novel hypotheses tailored for teleost species, without the need to rely on the limited number of fish-specific antibodies and pre-selected markers. Several recent studies on single-cell transcriptomes of fish have already identified previously unnoticed expression signatures and provided astonishing insights into the diversity of teleost leukocytes and the evolution of vertebrate immunity. Without a doubt, scRNA-seq in tandem with bioinformatics tools and state-of-the-art methods, will facilitate studying the teleost immune system by not only defining key markers, but also teaching us about lymphoid tissue organization, development/differentiation, cell-cell interactions, antigen receptor repertoires, states of health and disease, all across time and space in fishes. These advances will invite more researchers to develop the tools necessary to explore the immunology of fishes, which remain non-conventional animal models from which we have much to learn.


Subject(s)
Fishes/genetics , Fishes/immunology , Leukocytes/immunology , Leukocytes/metabolism , RNA-Seq , Single-Cell Analysis , Animals , Immunity , Single-Cell Analysis/methods
15.
Genes (Basel) ; 13(3)2022 03 15.
Article in English | MEDLINE | ID: covidwho-1760489

ABSTRACT

Despite the importance of and current demand for abaca (Musa textilis Nee) fiber, there has been limited study that capitalizes on RNA-seq to identify candidate genes associated with high fiber quality and bunchy top virus (AbBTV) resistance. Three varieties (Abuab, Inosa, and Tangongon), one wild banana variety (Musa balbisiana Colla) Pacol, and two developed backcrosses (Abuab × Pacol BC2 and BC3) were grown at the Institute of Plant Breeding (IPB), Laguna, Philippines. The pseudostems of 3-month-old suckers of each genotype were sampled for RNA-seq. Datasets were analyzed for differential expression (DE) implementing various model frameworks, including pairwise, genotypic and non-DE models. Results indicate that Abuab and BC3 induce the highest proportion (70%) of abaca-specific genes. Gene ontology (GO) enrichment analysis showed several genes associated with cellulose synthase activity, callose synthase, ß-glucosidase activity, glucan biosynthetic process, etc. KEGG pathway analysis showed several genes encoding for enzymes involved in the lignin biosynthetic pathway. Analysis using genotypic DE (GDE) between abaca bunchy top virus (AbBTV)-resistant and -susceptible groups revealed genes such as pathogenesis-related protein and NBS-LRR. As the genotypes were not infected with the pathogen, these genes are yet to be confirmed for their roles in disease resistance and are an interesting subject for further investigation.


Subject(s)
Musa , Carbohydrates , Dietary Fiber , Disease Resistance/genetics , Musa/genetics , Plant Breeding , RNA-Seq
16.
Int J Mol Sci ; 23(5)2022 Feb 24.
Article in English | MEDLINE | ID: covidwho-1715406

ABSTRACT

To better understand the molecular basis of respiratory diseases of viral origin, high-throughput gene-expression data are frequently taken by means of DNA microarray or RNA-seq technology. Such data can also be useful to classify infected individuals by molecular signatures in the form of machine-learning models with genes as predictor variables. Early diagnosis of patients by molecular signatures could also contribute to better treatments. An approach that has rarely been considered for machine-learning models in the context of transcriptomics is data augmentation. For other data types it has been shown that augmentation can improve classification accuracy and prevent overfitting. Here, we compare three strategies for data augmentation of DNA microarray and RNA-seq data from two selected studies on respiratory diseases of viral origin. The first study involves samples of patients with either viral or bacterial origin of the respiratory disease, the second study involves patients with either SARS-CoV-2 or another respiratory virus as disease origin. Specifically, we reanalyze these public datasets to study whether patient classification by transcriptomic signatures can be improved when adding artificial data for training of the machine-learning models. Our comparison reveals that augmentation of transcriptomic data can improve the classification accuracy and that fewer genes are necessary as explanatory variables in the final models. We also report genes from our signatures that overlap with signatures presented in the original publications of our example data. Due to strict selection criteria, the molecular role of these genes in the context of respiratory infectious diseases is underlined.


Subject(s)
COVID-19/genetics , Gene Expression Profiling/methods , Machine Learning , Neural Networks, Computer , RNA-Seq/methods , Transcriptome/genetics , Algorithms , COVID-19/classification , COVID-19/virology , Gene Ontology , Humans , Reproducibility of Results , SARS-CoV-2/physiology
17.
Sci Immunol ; 7(68): eabf2846, 2022 02 11.
Article in English | MEDLINE | ID: covidwho-1685480

ABSTRACT

Macrophages regulate protective immune responses to infectious microbes, but aberrant macrophage activation frequently drives pathological inflammation. To identify regulators of vigorous macrophage activation, we analyzed RNA-seq data from synovial macrophages and identified SLAMF7 as a receptor associated with a superactivated macrophage state in rheumatoid arthritis. We implicated IFN-γ as a key regulator of SLAMF7 expression and engaging SLAMF7 drove a strong wave of inflammatory cytokine expression. Induction of TNF-α after SLAMF7 engagement amplified inflammation through an autocrine signaling loop. We observed SLAMF7-induced gene programs not only in macrophages from rheumatoid arthritis patients but also in gut macrophages from patients with active Crohn's disease and in lung macrophages from patients with severe COVID-19. This suggests a central role for SLAMF7 in macrophage superactivation with broad implications in human disease pathology.


Subject(s)
Inflammation/immunology , Macrophage Activation/immunology , Signaling Lymphocytic Activation Molecule Family/immunology , Transcriptome/immunology , Acute Disease , Adult , Arthritis, Rheumatoid/genetics , Arthritis, Rheumatoid/immunology , Arthritis, Rheumatoid/metabolism , COVID-19/genetics , COVID-19/immunology , COVID-19/metabolism , COVID-19/virology , Cells, Cultured , Chronic Disease , Crohn Disease/genetics , Crohn Disease/immunology , Crohn Disease/metabolism , Female , Humans , Inflammation/genetics , Inflammation/metabolism , Macrophage Activation/genetics , RNA-Seq/methods , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2/immunology , SARS-CoV-2/physiology , Signaling Lymphocytic Activation Molecule Family/genetics , Signaling Lymphocytic Activation Molecule Family/metabolism , Single-Cell Analysis/methods , Synovial Membrane/immunology , Synovial Membrane/metabolism , Synovial Membrane/pathology , Transcriptome/genetics
18.
Database (Oxford) ; 20222022 01 17.
Article in English | MEDLINE | ID: covidwho-1672169

ABSTRACT

In recent years, efficient scRNA-seq methods have been developed, enabling the transcriptome profiling of single cells massively in parallel. Meanwhile, its high dimensionality and complexity bring challenges to the data analysis and require extensive collaborations between biologists and bioinformaticians and/or biostatisticians. The communication between these two units demands a platform for easy data sharing and exploration. Here we developed Single-Cell Transcriptomics Annotated Viewer (SCANNER), as a public web resource for the scientific community, for sharing and analyzing scRNA-seq data in a collaborative manner. It is easy-to-use without requiring special software or extensive coding skills. Moreover, it equipped a real-time database for secure data management and enables an efficient investigation of the activation of gene sets on a single-cell basis. Currently, SCANNER hosts a database of 19 types of cancers and COVID-19, as well as healthy samples from lungs of smokers and non-smokers, human brain cells and peripheral blood mononuclear cells (PBMC). The database will be frequently updated with datasets from new studies. Using SCANNER, we identified a larger proportion of cancer-associated fibroblasts cells and more active fibroblast growth-related genes in melanoma tissues in female patients compared to male patients. Moreover, we found ACE2 is mainly expressed in lung pneumocytes, secretory cells and ciliated cells and differentially expressed in lungs of smokers and never smokers.


Subject(s)
COVID-19 , Leukocytes, Mononuclear , Female , Gene Expression Profiling , Humans , Male , RNA-Seq , SARS-CoV-2 , Sequence Analysis, RNA , Single-Cell Analysis , Software
19.
Nature ; 602(7896): 268-273, 2022 02.
Article in English | MEDLINE | ID: covidwho-1671587

ABSTRACT

Genetic risk for autism spectrum disorder (ASD) is associated with hundreds of genes spanning a wide range of biological functions1-6. The alterations in the human brain resulting from mutations in these genes remain unclear. Furthermore, their phenotypic manifestation varies across individuals7,8. Here we used organoid models of the human cerebral cortex to identify cell-type-specific developmental abnormalities that result from haploinsufficiency in three ASD risk genes-SUV420H1 (also known as KMT5B), ARID1B and CHD8-in multiple cell lines from different donors, using single-cell RNA-sequencing (scRNA-seq) analysis of more than 745,000 cells and proteomic analysis of individual organoids, to identify phenotypic convergence. Each of the three mutations confers asynchronous development of two main cortical neuronal lineages-γ-aminobutyric-acid-releasing (GABAergic) neurons and deep-layer excitatory projection neurons-but acts through largely distinct molecular pathways. Although these phenotypes are consistent across cell lines, their expressivity is influenced by the individual genomic context, in a manner that is dependent on both the risk gene and the developmental defect. Calcium imaging in intact organoids shows that these early-stage developmental changes are followed by abnormal circuit activity. This research uncovers cell-type-specific neurodevelopmental abnormalities that are shared across ASD risk genes and are finely modulated by human genomic context, finding convergence in the neurobiological basis of how different risk genes contribute to ASD pathology.


Subject(s)
Autism Spectrum Disorder , Genetic Predisposition to Disease , Neurons , Autism Spectrum Disorder/genetics , Autism Spectrum Disorder/metabolism , Autism Spectrum Disorder/pathology , Cerebral Cortex/cytology , DNA-Binding Proteins/genetics , GABAergic Neurons/metabolism , GABAergic Neurons/pathology , Histone-Lysine N-Methyltransferase/genetics , Humans , Neurons/classification , Neurons/metabolism , Neurons/pathology , Organoids/cytology , Proteomics , RNA-Seq , Single-Cell Analysis , Transcription Factors/genetics
20.
Cells ; 11(3)2022 01 30.
Article in English | MEDLINE | ID: covidwho-1667057

ABSTRACT

The global outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is still ongoing, as is research on the molecular mechanisms underlying cellular infection by coronaviruses, with the hope of developing therapeutic agents against this pandemic. Other important respiratory viruses such as 2009 pandemic H1N1 and H7N9 avian influenza virus (AIV), influenza A viruses, are also responsible for a possible outbreak due to their respiratory susceptibility. However, the interaction of these viruses with host cells and the regulation of post-transcriptional genes remains unclear. In this study, we detected and analyzed the comparative transcriptome profiling of SARS-CoV-2, panH1N1 (A/California/07/2009), and H7N9 (A/Shanghai/1/2013) infected cells. The results showed that the commonly upregulated genes among the three groups were mainly involved in autophagy, pertussis, and tuberculosis, which indicated that autophagy plays an important role in viral pathogenicity. There are three groups of commonly downregulated genes involved in metabolic pathways. Notably, unlike panH1N1 and H7N9, SARS-CoV-2 infection can inhibit the m-TOR pathway and activate the p53 signaling pathway, which may be responsible for unique autophagy induction and cell apoptosis. Particularly, upregulated expression of IRF1 was found in SARS-CoV-2, panH1N1, and H7N9 infection. Further analysis showed SARS-CoV-2, panH1N1, and H7N9 infection-induced upregulation of lncRNA-34087.27 could serve as a competitive endogenous RNA to stabilize IRF1 mRNA by competitively binding with miR-302b-3p. This study provides new insights into the molecular mechanisms of influenza A virus and SARS-CoV-2 infection.


Subject(s)
COVID-19/immunology , Immunity/immunology , Influenza A Virus, H1N1 Subtype/immunology , Influenza A Virus, H7N9 Subtype/immunology , Influenza, Human/immunology , RNA/immunology , Transcriptome/immunology , A549 Cells , Animals , COVID-19/genetics , COVID-19/virology , HEK293 Cells , Host-Pathogen Interactions/immunology , Humans , Immunity/genetics , Influenza A Virus, H1N1 Subtype/physiology , Influenza A Virus, H7N9 Subtype/physiology , Influenza, Human/genetics , Influenza, Human/virology , Interferon Regulatory Factor-1/genetics , Interferon Regulatory Factor-1/immunology , Interferon Regulatory Factor-1/metabolism , MicroRNAs/genetics , MicroRNAs/immunology , MicroRNAs/metabolism , Pandemics/prevention & control , RNA/genetics , RNA/metabolism , RNA, Long Noncoding/genetics , RNA, Long Noncoding/immunology , RNA, Long Noncoding/metabolism , RNA, Messenger/genetics , RNA, Messenger/immunology , RNA, Messenger/metabolism , RNA-Seq/methods , SARS-CoV-2/physiology , Signal Transduction/genetics , Signal Transduction/immunology , Transcriptome/genetics
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