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COVID-19, which is caused by SARS-CoV-2, can result in acute respiratory distress syndrome and multiple organ failure1-4, but little is known about its pathophysiology. Here we generated single-cell atlases of 24 lung, 16 kidney, 16 liver and 19 heart autopsy tissue samples and spatial atlases of 14 lung samples from donors who died of COVID-19. Integrated computational analysis uncovered substantial remodelling in the lung epithelial, immune and stromal compartments, with evidence of multiple paths of failed tissue regeneration, including defective alveolar type 2 differentiation and expansion of fibroblasts and putative TP63+ intrapulmonary basal-like progenitor cells. Viral RNAs were enriched in mononuclear phagocytic and endothelial lung cells, which induced specific host programs. Spatial analysis in lung distinguished inflammatory host responses in lung regions with and without viral RNA. Analysis of the other tissue atlases showed transcriptional alterations in multiple cell types in heart tissue from donors with COVID-19, and mapped cell types and genes implicated with disease severity based on COVID-19 genome-wide association studies. Our foundational dataset elucidates the biological effect of severe SARS-CoV-2 infection across the body, a key step towards new treatments.
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COVID-19/patología , COVID-19/virología , Riñón/patología , Hígado/patología , Pulmón/patología , Miocardio/patología , SARS-CoV-2/patogenicidad , Adulto , Anciano , Anciano de 80 o más Años , Atlas como Asunto , Autopsia , Bancos de Muestras Biológicas , COVID-19/genética , COVID-19/inmunología , Células Endoteliales , Células Epiteliales/patología , Células Epiteliales/virología , Femenino , Fibroblastos , Estudio de Asociación del Genoma Completo , Corazón/virología , Humanos , Inflamación/patología , Inflamación/virología , Riñón/virología , Hígado/virología , Pulmón/virología , Masculino , Persona de Mediana Edad , Especificidad de Órganos , Fagocitos , Alveolos Pulmonares/patología , Alveolos Pulmonares/virología , ARN Viral/análisis , Regeneración , SARS-CoV-2/inmunología , Análisis de la Célula Individual , Carga ViralRESUMEN
Mammalian gene expression is inherently stochastic1,2, and results in discrete bursts of RNA molecules that are synthesized from each allele3-7. Although transcription is known to be regulated by promoters and enhancers, it is unclear how cis-regulatory sequences encode transcriptional burst kinetics. Characterization of transcriptional bursting, including the burst size and frequency, has mainly relied on live-cell4,6,8 or single-molecule RNA fluorescence in situ hybridization3,5,8,9 recordings of selected loci. Here we determine transcriptome-wide burst frequencies and sizes for endogenous mouse and human genes using allele-sensitive single-cell RNA sequencing. We show that core promoter elements affect burst size and uncover synergistic effects between TATA and initiator elements, which were masked at mean expression levels. Notably, we provide transcriptome-wide evidence that enhancers control burst frequencies, and demonstrate that cell-type-specific gene expression is primarily shaped by changes in burst frequencies. Together, our data show that burst frequency is primarily encoded in enhancers and burst size in core promoters, and that allelic single-cell RNA sequencing is a powerful model for investigating transcriptional kinetics.
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Genes/genética , Genómica , Transcripción Genética/genética , Alelos , Animales , Elementos de Facilitación Genéticos/genética , Fibroblastos/metabolismo , Humanos , Cinética , Masculino , Ratones , Células Madre Embrionarias de Ratones/metabolismo , Especificidad de Órganos/genética , Polimorfismo Genético , Regiones Promotoras Genéticas/genética , Análisis de Secuencia de ARN , Eliminación de Secuencia , Análisis de la Célula Individual , Procesos Estocásticos , TATA Box/genética , Transcriptoma/genéticaRESUMEN
Charting an organs' biological atlas requires us to spatially resolve the entire single-cell transcriptome, and to relate such cellular features to the anatomical scale. Single-cell and single-nucleus RNA-seq (sc/snRNA-seq) can profile cells comprehensively, but lose spatial information. Spatial transcriptomics allows for spatial measurements, but at lower resolution and with limited sensitivity. Targeted in situ technologies solve both issues, but are limited in gene throughput. To overcome these limitations we present Tangram, a method that aligns sc/snRNA-seq data to various forms of spatial data collected from the same region, including MERFISH, STARmap, smFISH, Spatial Transcriptomics (Visium) and histological images. Tangram can map any type of sc/snRNA-seq data, including multimodal data such as those from SHARE-seq, which we used to reveal spatial patterns of chromatin accessibility. We demonstrate Tangram on healthy mouse brain tissue, by reconstructing a genome-wide anatomically integrated spatial map at single-cell resolution of the visual and somatomotor areas.
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Encéfalo/metabolismo , Cromatina/genética , Aprendizaje Profundo , Regulación de la Expresión Génica , Análisis de la Célula Individual/métodos , Programas Informáticos , Transcriptoma , Animales , Cromatina/química , Cromatina/metabolismo , Femenino , Perfilación de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , RNA-Seq , Secuencias Reguladoras de Ácidos NucleicosRESUMEN
Ca2+-sensor proteins are generally implicated in insulin release through SNARE interactions. Here, secretagogin, whose expression in human pancreatic islets correlates with their insulin content and the incidence of type 2 diabetes, is shown to orchestrate an unexpectedly distinct mechanism. Single-cell RNA-seq reveals retained expression of the TRP family members in ß-cells from diabetic donors. Amongst these, pharmacological probing identifies Ca2+-permeable transient receptor potential vanilloid type 1 channels (TRPV1) as potent inducers of secretagogin expression through recruitment of Sp1 transcription factors. Accordingly, agonist stimulation of TRPV1s fails to rescue insulin release from pancreatic islets of glucose intolerant secretagogin knock-out(-/-) mice. However, instead of merely impinging on the SNARE machinery, reduced insulin availability in secretagogin-/- mice is due to ß-cell loss, which is underpinned by the collapse of protein folding and deregulation of secretagogin-dependent USP9X deubiquitinase activity. Therefore, and considering the desensitization of TRPV1s in diabetic pancreata, a TRPV1-to-secretagogin regulatory axis seems critical to maintain the structural integrity and signal competence of ß-cells.
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Regulación de la Expresión Génica , Células Secretoras de Insulina/fisiología , Proteínas/metabolismo , Secretagoginas/metabolismo , Canales Catiónicos TRPV/metabolismo , Animales , Supervivencia Celular , Perfilación de la Expresión Génica , Humanos , Ratones , Ratones Noqueados , Secretagoginas/deficiencia , Análisis de la Célula IndividualRESUMEN
In vivo UV crosslinking identified numerous preribosomal RNA (pre-rRNA) binding sites for the large, highly conserved ribosome synthesis factor Rrp5. Intramolecular complementation has shown that the C-terminal domain (CTD) of Rrp5 is required for pre-rRNA cleavage at sites A0-A2 on the pathway of 18S rRNA synthesis, whereas the N-terminal domain (NTD) is required for A3 cleavage on the pathway of 5.8S/25S rRNA synthesis. The CTD was crosslinked to sequences flanking A2 and to the snoRNAs U3, U14, snR30, and snR10, which are required for cleavage at A0-A2. The NTD was crosslinked to sequences flanking A3 and to the RNA component of ribonuclease MRP, which cleaves site A3. Rrp5 could also be directly crosslinked to several large structural proteins and nucleoside triphosphatases. A key role in coordinating preribosomal assembly and processing was confirmed by chromatin spreads. Following depletion of Rrp5, cotranscriptional cleavage was lost and preribosome compaction greatly reduced.
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Proteínas Fúngicas/genética , Precursores del ARN/genética , Procesamiento Postranscripcional del ARN , ARN de Hongos/genética , ARN Ribosómico/genética , Ribosomas/genética , Secuencia de Bases , Sitios de Unión , Endorribonucleasas/genética , Endorribonucleasas/metabolismo , Proteínas Fúngicas/metabolismo , Datos de Secuencia Molecular , Nucleósido-Trifosfatasa/genética , Nucleósido-Trifosfatasa/metabolismo , Precursores del ARN/metabolismo , ARN Ribosómico/metabolismo , Levaduras/genética , Levaduras/metabolismoRESUMEN
Ribosomal subunit biogenesis in eukaryotes is a complex multistep process. Mrd1 is an essential and conserved small (40S) ribosomal subunit synthesis factor that is required for early cleavages in the 35S pre-ribosomal RNA (rRNA). Yeast Mrd1 contains five RNA-binding domains (RBDs), all of which are necessary for optimal function of the protein. Proteomic data showed that Mrd1 is part of the early pre-ribosomal complexes, and deletion of individual RBDs perturbs the pre-ribosomal structure. In vivo ultraviolet cross-linking showed that Mrd1 binds to the pre-rRNA at two sites within the 18S region, in helix 27 (h27) and helix 28. The major binding site lies in h27, and mutational analyses shows that this interaction requires the RBD1-3 region of Mrd1. RBD2 plays the dominant role in h27 binding, but other RBDs also contribute directly. h27 and helix 28 are located close to the sequences that form the central pseudoknot, a key structural feature of the mature 40S subunit. We speculate that the modular structure of Mrd1 coordinates pseudoknot formation with pre-rRNA processing and subunit assembly.
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Precursores del ARN/metabolismo , ARN Ribosómico 18S/metabolismo , Proteínas de Unión al ARN/metabolismo , Subunidades Ribosómicas Pequeñas de Eucariotas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Sitios de Unión , Conformación de Ácido Nucleico , Estructura Terciaria de Proteína , Precursores del ARN/química , ARN Ribosómico 18S/química , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/genética , Subunidades Ribosómicas Pequeñas de Eucariotas/química , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Eliminación de SecuenciaRESUMEN
Population genetics continues to identify genetic variants associated with diseases of the immune system and offers a unique opportunity to discover mechanisms of immune regulation. Multiple genetic variants linked to severe fungal infections and autoimmunity are associated with caspase recruitment domain-containing protein 9 (CARD9). We leverage the CARD9 R101C missense variant to uncover a biochemical mechanism of CARD9 activation essential for antifungal responses. We demonstrate that R101C disrupts a critical signaling switch whereby phosphorylation of S104 releases CARD9 from an autoinhibited state to promote inflammatory responses in myeloid cells. Furthermore, we show that CARD9 R101C exerts dynamic effects on the skin cellular contexture during fungal infection, corrupting inflammatory signaling and cell-cell communication circuits. Card9 R101C mice fail to control dermatophyte infection in the skin, resulting in high fungal burden, yet show minimal signs of inflammation. Together, we demonstrate how translational genetics reveals molecular and cellular mechanisms of innate immune regulation.
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Proteínas Adaptadoras de Señalización CARD , Micosis , Animales , Ratones , Fosforilación , Proteínas Adaptadoras de Señalización CARD/metabolismo , Transducción de Señal , Inflamación , AntifúngicosRESUMEN
TP53 is the most frequently mutated gene across many cancers and is associated with shorter survival in lung adenocarcinoma (LUAD). To define how TP53 mutations affect the LUAD tumor microenvironment (TME), we constructed a multi-omic cellular and spatial tumor atlas of 23 treatment-naïve human lung tumors. We found that TP53 -mutant ( TP53 mut ) malignant cells lose alveolar identity and upregulate highly proliferative and entropic gene expression programs consistently across resectable LUAD patient tumors, genetically engineered mouse models, and cell lines harboring a wide spectrum of TP53 mutations. We further identified a multicellular tumor niche composed of SPP1 + macrophages and collagen-expressing fibroblasts that coincides with hypoxic, pro-metastatic expression programs in TP53 mut tumors. Spatially correlated angiostatic and immune checkpoint interactions, including CD274 - PDCD1 and PVR - TIGIT , are also enriched in TP53 mut LUAD tumors, which may influence response to checkpoint blockade therapy. Our methodology can be further applied to investigate mutation-specific TME changes in other cancers.
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Cardiovascular disease plays a central role in the electrical and structural remodeling of the right atrium, predisposing to arrhythmias, heart failure, and sudden death. Here, we dissect with single-nuclei RNA sequencing (snRNA-seq) and spatial transcriptomics the gene expression changes in the human ex vivo right atrial tissue and pericardial fluid in ischemic heart disease, myocardial infarction, and ischemic and non-ischemic heart failure using asymptomatic patients with valvular disease who undergo preventive surgery as the control group. We reveal substantial differences in disease-associated gene expression in all cell types, collectively suggesting inflammatory microvascular dysfunction and changes in the right atrial tissue composition as the valvular and vascular diseases progress into heart failure. The data collectively suggest that investigation of human cardiovascular disease should expand to all functionally important parts of the heart, which may help us to identify mechanisms promoting more severe types of the disease.
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Atrios Cardíacos , Microvasos , Isquemia Miocárdica , Transcriptoma , Humanos , Atrios Cardíacos/patología , Atrios Cardíacos/metabolismo , Isquemia Miocárdica/genética , Isquemia Miocárdica/patología , Isquemia Miocárdica/metabolismo , Transcriptoma/genética , Microvasos/patología , Inflamación/patología , Inflamación/genética , Masculino , Femenino , Persona de Mediana Edad , Anciano , Regulación de la Expresión GénicaRESUMEN
Neuroblastoma is a pediatric cancer arising from the developing sympathoadrenal lineage with complex inter- and intra-tumoral heterogeneity. To chart this complexity, we generated a comprehensive cell atlas of 55 neuroblastoma patient tumors, collected from two pediatric cancer institutions, spanning a range of clinical, genetic, and histologic features. Our atlas combines single-cell/nucleus RNA-seq (sc/scRNA-seq), bulk RNA-seq, whole exome sequencing, DNA methylation profiling, spatial transcriptomics, and two spatial proteomic methods. Sc/snRNA-seq revealed three malignant cell states with features of sympathoadrenal lineage development. All of the neuroblastomas had malignant cells that resembled sympathoblasts and the more differentiated adrenergic cells. A subset of tumors had malignant cells in a mesenchymal cell state with molecular features of Schwann cell precursors. DNA methylation profiles defined four groupings of patients, which differ in the degree of malignant cell heterogeneity and clinical outcomes. Using spatial proteomics, we found that neuroblastomas are spatially compartmentalized, with malignant tumor cells sequestered away from immune cells. Finally, we identify spatially restricted signaling patterns in immune cells from spatial transcriptomics. To facilitate the visualization and analysis of our atlas as a resource for further research in neuroblastoma, single cell, and spatial-omics, all data are shared through the Human Tumor Atlas Network Data Commons at www.humantumoratlas.org.
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Building a blastema from the stump is a key step of salamander limb regeneration. Stump-derived cells temporarily suspend their identity as they contribute to the blastema by a process generally referred to as dedifferentiation. Here, we provide evidence for a mechanism that involves an active inhibition of protein synthesis during blastema formation and growth. Relieving this inhibition results in a higher number of cycling cells and enhances the pace of limb regeneration. By small RNA profiling and fate mapping of skeletal muscle progeny as a cellular model for dedifferentiation, we find that the downregulation of miR-10b-5p is critical for rebooting the translation machinery. miR-10b-5p targets ribosomal mRNAs, and its artificial upregulation causes decreased blastema cell proliferation, reduction in transcripts that encode ribosomal subunits, diminished nascent protein synthesis, and retardation of limb regeneration. Taken together, our data identify a link between miRNA regulation, ribosome biogenesis, and protein synthesis during newt limb regeneration.
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MicroARNs , ARN Pequeño no Traducido , Animales , Urodelos/genética , ARN Pequeño no Traducido/metabolismo , Músculo Esquelético/metabolismo , Ribosomas/genética , MicroARNs/genética , MicroARNs/metabolismo , Extremidades/fisiologíaRESUMEN
Effective tissue repair requires coordinated intercellular communication to sense damage, remodel the tissue, and restore function. Here, we dissected the healing response in the intestinal mucosa by mapping intercellular communication at single-cell resolution and integrating with spatial transcriptomics. We demonstrated that a risk variant for Crohn's disease, hepatocyte growth factor activator (HGFAC) Arg509His (R509H), disrupted a damage-sensing pathway connecting the coagulation cascade to growth factors that drive the differentiation of wound-associated epithelial (WAE) cells and production of a localized retinoic acid (RA) gradient to promote fibroblast-mediated tissue remodeling. Specifically, we showed that HGFAC R509H was activated by thrombin protease activity but exhibited impaired proteolytic activation of the growth factor macrophage-stimulating protein (MSP). In Hgfac R509H mice, reduced MSP activation in response to wounding of the colon resulted in impaired WAE cell induction and delayed healing. Through integration of single-cell transcriptomics and spatial transcriptomics, we demonstrated that WAE cells generated RA in a spatially restricted region of the wound site and that mucosal fibroblasts responded to this signal by producing extracellular matrix and growth factors. We further dissected this WAE cell-fibroblast signaling circuit in vitro using a genetically tractable organoid coculture model. Collectively, these studies exploited a genetic perturbation associated with human disease to disrupt a fundamental biological process and then reconstructed a spatially resolved mechanistic model of tissue healing.
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Enfermedad de Crohn , Ratones , Humanos , Animales , Enfermedad de Crohn/genética , Enfermedad de Crohn/metabolismo , Transducción de Señal , Células Epiteliales/metabolismo , Mucosa Intestinal/metabolismo , Diferenciación CelularRESUMEN
Tissue biology involves an intricate balance between cell-intrinsic processes and interactions between cells organized in specific spatial patterns, which can be respectively captured by single-cell profiling methods, such as single-cell RNA-seq (scRNA-seq), and histology imaging data, such as Hematoxylin-and-Eosin (H&E) stains. While single-cell profiles provide rich molecular information, they can be challenging to collect routinely and do not have spatial resolution. Conversely, histological H&E assays have been a cornerstone of tissue pathology for decades, but do not directly report on molecular details, although the observed structure they capture arises from molecules and cells. Here, we leverage adversarial machine learning to develop SCHAF (Single-Cell omics from Histology Analysis Framework), to generate a tissue sample's spatially-resolved single-cell omics dataset from its H&E histology image. We demonstrate SCHAF on two types of human tumors-from lung and metastatic breast cancer-training with matched samples analyzed by both sc/snRNA-seq and by H&E staining. SCHAF generated appropriate single-cell profiles from histology images in test data, related them spatially, and compared well to ground-truth scRNA-Seq, expert pathologist annotations, or direct MERFISH measurements. SCHAF opens the way to next-generation H&E2.0 analyses and an integrated understanding of cell and tissue biology in health and disease.
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The critical functions of the human liver are coordinated through the interactions of hepatic parenchymal and non-parenchymal cells. Recent advances in single-cell transcriptional approaches have enabled an examination of the human liver with unprecedented resolution. However, dissociation-related cell perturbation can limit the ability to fully capture the human liver's parenchymal cell fraction, which limits the ability to comprehensively profile this organ. Here, we report the transcriptional landscape of 73,295 cells from the human liver using matched single-cell RNA sequencing (scRNA-seq) and single-nucleus RNA sequencing (snRNA-seq). The addition of snRNA-seq enabled the characterization of interzonal hepatocytes at a single-cell resolution, revealed the presence of rare subtypes of liver mesenchymal cells, and facilitated the detection of cholangiocyte progenitors that had only been observed during in vitro differentiation experiments. However, T and B lymphocytes and natural killer cells were only distinguishable using scRNA-seq, highlighting the importance of applying both technologies to obtain a complete map of tissue-resident cell types. We validated the distinct spatial distribution of the hepatocyte, cholangiocyte, and mesenchymal cell populations by an independent spatial transcriptomics data set and immunohistochemistry. Conclusion: Our study provides a systematic comparison of the transcriptomes captured by scRNA-seq and snRNA-seq and delivers a high-resolution map of the parenchymal cell populations in the healthy human liver.
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Hígado , Análisis de la Célula Individual , Núcleo Celular/genética , Humanos , Análisis de Secuencia de ARN , Transcriptoma/genéticaRESUMEN
Synovial sarcoma (SyS) is an aggressive neoplasm driven by the SS18-SSX fusion, and is characterized by low T cell infiltration. Here, we studied the cancer-immune interplay in SyS using an integrative approach that combines single-cell RNA sequencing (scRNA-seq), spatial profiling and genetic and pharmacological perturbations. scRNA-seq of 16,872 cells from 12 human SyS tumors uncovered a malignant subpopulation that marks immune-deprived niches in situ and is predictive of poor clinical outcomes in two independent cohorts. Functional analyses revealed that this malignant cell state is controlled by the SS18-SSX fusion, is repressed by cytokines secreted by macrophages and T cells, and can be synergistically targeted with a combination of HDAC and CDK4/CDK6 inhibitors. This drug combination enhanced malignant-cell immunogenicity in SyS models, leading to induced T cell reactivity and T cell-mediated killing. Our study provides a blueprint for investigating heterogeneity in fusion-driven malignancies and demonstrates an interplay between immune evasion and oncogenic processes that can be co-targeted in SyS and potentially in other malignancies.
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Carcinogénesis/genética , Terapia Molecular Dirigida , Proteínas de Fusión Oncogénica/genética , Sarcoma Sinovial/tratamiento farmacológico , Línea Celular Tumoral , Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Inhibidores de Histona Desacetilasas/uso terapéutico , Histona Desacetilasas/genética , Histona Desacetilasas/uso terapéutico , Humanos , Proteínas de Fusión Oncogénica/antagonistas & inhibidores , Oncogenes/genética , RNA-Seq , Sarcoma Sinovial/genética , Sarcoma Sinovial/patología , Análisis de la Célula IndividualRESUMEN
The SARS-CoV-2 pandemic has caused over 1 million deaths globally, mostly due to acute lung injury and acute respiratory distress syndrome, or direct complications resulting in multiple-organ failures. Little is known about the host tissue immune and cellular responses associated with COVID-19 infection, symptoms, and lethality. To address this, we collected tissues from 11 organs during the clinical autopsy of 17 individuals who succumbed to COVID-19, resulting in a tissue bank of approximately 420 specimens. We generated comprehensive cellular maps capturing COVID-19 biology related to patients' demise through single-cell and single-nucleus RNA-Seq of lung, kidney, liver and heart tissues, and further contextualized our findings through spatial RNA profiling of distinct lung regions. We developed a computational framework that incorporates removal of ambient RNA and automated cell type annotation to facilitate comparison with other healthy and diseased tissue atlases. In the lung, we uncovered significantly altered transcriptional programs within the epithelial, immune, and stromal compartments and cell intrinsic changes in multiple cell types relative to lung tissue from healthy controls. We observed evidence of: alveolar type 2 (AT2) differentiation replacing depleted alveolar type 1 (AT1) lung epithelial cells, as previously seen in fibrosis; a concomitant increase in myofibroblasts reflective of defective tissue repair; and, putative TP63+ intrapulmonary basal-like progenitor (IPBLP) cells, similar to cells identified in H1N1 influenza, that may serve as an emergency cellular reserve for severely damaged alveoli. Together, these findings suggest the activation and failure of multiple avenues for regeneration of the epithelium in these terminal lungs. SARS-CoV-2 RNA reads were enriched in lung mononuclear phagocytic cells and endothelial cells, and these cells expressed distinct host response transcriptional programs. We corroborated the compositional and transcriptional changes in lung tissue through spatial analysis of RNA profiles in situ and distinguished unique tissue host responses between regions with and without viral RNA, and in COVID-19 donor tissues relative to healthy lung. Finally, we analyzed genetic regions implicated in COVID-19 GWAS with transcriptomic data to implicate specific cell types and genes associated with disease severity. Overall, our COVID-19 cell atlas is a foundational dataset to better understand the biological impact of SARS-CoV-2 infection across the human body and empowers the identification of new therapeutic interventions and prevention strategies.
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In Saccharomyces cerevisiae, synthesis of the small ribosomal subunit requires assembly of the 35S pre-rRNA into a 90S preribosomal complex. SnoRNAs, including U3 snoRNA, and many trans-acting proteins are required for the ordered assembly and function of the 90S preribosomal complex. Here, we show that the conserved protein Mrd1p binds to the pre-rRNA early during transcription and is required for compaction of the pre-18S rRNA into SSU processome particles. We have exploited the fact that an Mrd1p-GFP fusion protein is incorporated into the 90S preribosomal complex, where it acts as a partial loss-of-function mutation. When associated with the pre-rRNA, Mrd1p-GFP functionally interacts with the essential Pwp2, Mpp10 and U3 snoRNP subcomplexes that are functionally interconnected in the 90S preribosomal complex. The fusion protein can partially support 90S preribosome-mediated cleavages at the A(0)-A(2) sites. At the same time, on a substantial fraction of transcripts, the composition and/or structure of the 90S preribosomal complex is perturbed by the fusion protein in such a way that cleavage of the 35S pre-rRNA is either blocked or shifted to aberrant sites. These results show that Mrd1p is required for establishing productive structures within the 90S preribosomal complex.
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Precursores del ARN/metabolismo , ARN Ribosómico/metabolismo , Proteínas de Unión al ARN/metabolismo , Subunidades Ribosómicas Pequeñas de Eucariotas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Nucléolo Celular/metabolismo , Núcleo Celular/metabolismo , Mutación , Fosfoproteínas/metabolismo , Procesamiento Postranscripcional del ARN , ARN Nucleolar Pequeño/metabolismo , Proteínas de Unión al ARN/genética , Proteínas Recombinantes de Fusión/metabolismo , Ribonucleoproteínas/metabolismo , Proteínas Ribosómicas , Subunidades Ribosómicas Pequeñas de Eucariotas/química , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Transcripción GenéticaRESUMEN
Many important cell types in adult vertebrates have a mesenchymal origin, including fibroblasts and vascular mural cells. Although their biological importance is undisputed, the level of mesenchymal cell heterogeneity within and between organs, while appreciated, has not been analyzed in detail. Here, we compare single-cell transcriptional profiles of fibroblasts and vascular mural cells across four murine muscular organs: heart, skeletal muscle, intestine and bladder. We reveal gene expression signatures that demarcate fibroblasts from mural cells and provide molecular signatures for cell subtype identification. We observe striking inter- and intra-organ heterogeneity amongst the fibroblasts, primarily reflecting differences in the expression of extracellular matrix components. Fibroblast subtypes localize to discrete anatomical positions offering novel predictions about physiological function(s) and regulatory signaling circuits. Our data shed new light on the diversity of poorly defined classes of cells and provide a foundation for improved understanding of their roles in physiological and pathological processes.
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Diferenciación Celular , Fibroblastos/fisiología , Células Madre Mesenquimatosas/fisiología , Miocitos del Músculo Liso/fisiología , Pericitos/fisiología , Animales , Separación Celular , Vasos Coronarios/citología , Matriz Extracelular/metabolismo , Fibroblastos/citología , Citometría de Flujo , Intestinos/irrigación sanguínea , Intestinos/citología , Masculino , Ratones , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/citología , Músculo Liso Vascular/citología , Miocardio/citología , Miocitos del Músculo Liso/citología , Pericitos/citología , RNA-Seq , Análisis de la Célula Individual , Vejiga Urinaria/irrigación sanguínea , Vejiga Urinaria/citologíaRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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Vascular diseases are major causes of death, yet our understanding of the cellular constituents of blood vessels, including how differences in their gene expression profiles create diversity in vascular structure and function, is limited. In this paper, we describe a single-cell RNA sequencing (scRNA-seq) dataset that defines vascular and vessel-associated cell types and subtypes in mouse brain and lung. The dataset contains 3,436 single cell transcriptomes from mouse brain, which formed 15 distinct clusters corresponding to cell (sub)types, and another 1,504 single cell transcriptomes from mouse lung, which formed 17 cell clusters. In order to allow user-friendly access to our data, we constructed a searchable database (http://betsholtzlab.org/VascularSingleCells/database.html). Our dataset constitutes a comprehensive molecular atlas of vascular and vessel-associated cell types in the mouse brain and lung, and as such provides a strong foundation for future studies of vascular development and diseases.