Project description:The RNA sequencing approach has been broadly used to provide gene-, pathway-, and network-centric analyses for various cell and tissue samples. However, thus far, rich cellular information carried in tissue samples has not been thoroughly characterized from RNA-Seq data. Therefore, it would expand our horizons to better understand the biological processes of the body by incorporating a cell-centric view of tissue transcriptome. Here, a computational model named seq-ImmuCC was developed to infer the relative proportions of 10 major immune cells in mouse tissues from RNA-Seq data. The performance of seq-ImmuCC was evaluated among multiple computational algorithms, transcriptional platforms, and simulated and experimental datasets. The test results showed its stable performance and superb consistency with experimental observations under different conditions. With seq-ImmuCC, we generated the comprehensive landscape of immune cell compositions in 27 normal mouse tissues and extracted the distinct signatures of immune cell proportion among various tissue types. Furthermore, we quantitatively characterized and compared 18 different types of mouse tumor tissues of distinct cell origins with their immune cell compositions, which provided a comprehensive and informative measurement for the immune microenvironment inside tumor tissues. The online server of seq-ImmuCC are freely available at http://wap-lab.org:3200/immune/.
Project description:Knowledge of cell type composition in disease relevant tissues is an important step towards the identification of cellular targets of disease. We present MuSiC, a method that utilizes cell-type specific gene expression from single-cell RNA sequencing (RNA-seq) data to characterize cell type compositions from bulk RNA-seq data in complex tissues. By appropriate weighting of genes showing cross-subject and cross-cell consistency, MuSiC enables the transfer of cell type-specific gene expression information from one dataset to another. When applied to pancreatic islet and whole kidney expression data in human, mouse, and rats, MuSiC outperformed existing methods, especially for tissues with closely related cell types. MuSiC enables the characterization of cellular heterogeneity of complex tissues for understanding of disease mechanisms. As bulk tissue data are more easily accessible than single-cell RNA-seq, MuSiC allows the utilization of the vast amounts of disease relevant bulk tissue RNA-seq data for elucidating cell type contributions in disease.
Project description:Damage-associated microglia (DAM) profiles observed in Alzheimer's disease (AD)-related mouse models reflect an activation state that could modulate AD risk or progression. To learn whether human AD microglia (HAM) display a similar profile, we develop a method for purifying cell types from frozen cerebrocortical tissues for RNA-seq analysis, allowing better transcriptome coverage than typical single-nucleus RNA-seq approaches. The HAM profile we observe bears little resemblance to the DAM profile. Instead, HAM display an enhanced human aging profile, in addition to other disease-related changes such as APOE upregulation. Analyses of whole-tissue RNA-seq and single-cell/nucleus RNA-seq datasets corroborate our findings and suggest that the lack of DAM response in human microglia occurs specifically in AD tissues, not other neurodegenerative settings. These results, which can be browsed at http://research-pub.gene.com/BrainMyeloidLandscape, provide a genome-wide picture of microglial activation in human AD and highlight considerable differences between mouse models and human disease.
Project description:Single-cell RNA-seq is a powerful tool in decoding the heterogeneity in complex tissues by generating transcriptomic profiles of the individual cell. Here, we report a single-nuclei RNA-seq (snRNA-seq) transcriptomic study on human retinal tissue, which is composed of multiple cell types with distinct functions. Six samples from three healthy donors are profiled and high-quality RNA-seq data is obtained for 5873 single nuclei. All major retinal cell types are observed and marker genes for each cell type are identified. The gene expression of the macular and peripheral retina is compared to each other at cell-type level. Furthermore, our dataset shows an improved power for prioritizing genes associated with human retinal diseases compared to both mouse single-cell RNA-seq and human bulk RNA-seq results. In conclusion, we demonstrate that obtaining single cell transcriptomes from human frozen tissues can provide insight missed by either human bulk RNA-seq or animal models.
Project description:Discovering and classifying long noncoding RNAs (lncRNAs) across all mammalian tissues and cell lines remains a major challenge. Previously, mouse lncRNAs were identified using transcriptome sequencing (RNA-seq) data from a limited number of tissues or cell lines. Additionally, associating a few hundred lncRNA promoters with chromatin states in a single mouse cell line has identified two classes of chromatin-associated lncRNA. However, the discovery and classification of lncRNAs is still pending in many other tissues in mouse. To address this, we built a comprehensive catalog of lncRNAs by combining known lncRNAs with high-confidence novel lncRNAs identified by mapping and de novo assembling billions of RNA-seq reads from eight tissues and a primary cell line in mouse. Next, we integrated this catalog of lncRNAs with multiple genome-wide chromatin state maps and found two different classes of chromatin state-associated lncRNAs, including promoter-associated (plncRNAs) and enhancer-associated (elncRNAs) lncRNAs, across various tissues. Experimental knockdown of an elncRNA resulted in the downregulation of the neighboring protein-coding Kdm8 gene, encoding a histone demethylase. Our findings provide 2,803 novel lncRNAs and a comprehensive catalog of chromatin-associated lncRNAs across different tissues in mouse.
Project description:Single-nucleus RNA sequencing (sNuc-seq) profiles RNA from tissues that are preserved or cannot be dissociated, but it does not provide high throughput. Here, we develop DroNc-seq: massively parallel sNuc-seq with droplet technology. We profile 39,111 nuclei from mouse and human archived brain samples to demonstrate sensitive, efficient, and unbiased classification of cell types, paving the way for systematic charting of cell atlases.
Project description:The human eye is built from several specialized tissues which direct, capture and pre-process information to provide vision. The gene expression of the different eye tissues has been extensively profiled with RNA-seq across numerous studies. Large consortium projects have also used RNA-seq to study gene expression patterning across many different human tissues, minus the eye. There has not been an integrated study of expression patterns from multiple eye tissues compared with other human body tissues. We have collated all publicly available healthy human eye RNA-seq datasets as well as dozens of other tissues. We use this fully integrated dataset to probe the biological processes and pan expression relationships between the cornea, retina, retinal pigment epithelium (RPE)-choroid complex, and the rest of the human tissues with differential expression, clustering and gene ontology term enrichment tools. We also leverage our large collection of retina and RPE-choroid tissues to build the first human weighted gene correlation networks and use them to highlight known biological pathways and eye gene disease enrichment. We also have integrated publicly available single-cell RNA-seq data from mouse retina into our framework for validation and discovery. Finally, we make all these data, analyses and visualizations available via a powerful interactive web application (https://eyeintegration.nei.nih.gov/).
Project description:Transcriptomic profiling of complex tissues by single-nucleus RNA-sequencing (snRNA-seq) affords some advantages over single-cell RNA-sequencing (scRNA-seq). snRNA-seq provides less biased cellular coverage, does not appear to suffer cell isolation-based transcriptional artifacts, and can be applied to archived frozen specimens. We used well-matched snRNA-seq and scRNA-seq datasets from mouse visual cortex to compare cell type detection. Although more transcripts are detected in individual whole cells (~11,000 genes) than nuclei (~7,000 genes), we demonstrate that closely related neuronal cell types can be similarly discriminated with both methods if intronic sequences are included in snRNA-seq analysis. We estimate that the nuclear proportion of total cellular mRNA varies from 20% to over 50% for large and small pyramidal neurons, respectively. Together, these results illustrate the high information content of nuclear RNA for characterization of cellular diversity in brain tissues.
Project description:RNA sequencing (RNA-seq) is the leading technology for genome-wide transcript quantification. However, publicly available RNA-seq data is currently provided mostly in raw form, a significant barrier for global and integrative retrospective analyses. ARCHS4 is a web resource that makes the majority of published RNA-seq data from human and mouse available at the gene and transcript levels. For developing ARCHS4, available FASTQ files from RNA-seq experiments from the Gene Expression Omnibus (GEO) were aligned using a cloud-based infrastructure. In total 187,946 samples are accessible through ARCHS4 with 103,083 mouse and 84,863 human. Additionally, the ARCHS4 web interface provides intuitive exploration of the processed data through querying tools, interactive visualization, and gene pages that provide average expression across cell lines and tissues, top co-expressed genes for each gene, and predicted biological functions and protein-protein interactions for each gene based on prior knowledge combined with co-expression.
Project description:Spatial positions of cells in tissues strongly influence function, yet a high-throughput, genome-wide readout of gene expression with cellular resolution is lacking. We developed Slide-seq, a method for transferring RNA from tissue sections onto a surface covered in DNA-barcoded beads with known positions, allowing the locations of the RNA to be inferred by sequencing. Using Slide-seq, we localized cell types identified by single-cell RNA sequencing datasets within the cerebellum and hippocampus, characterized spatial gene expression patterns in the Purkinje layer of mouse cerebellum, and defined the temporal evolution of cell type-specific responses in a mouse model of traumatic brain injury. These studies highlight how Slide-seq provides a scalable method for obtaining spatially resolved gene expression data at resolutions comparable to the sizes of individual cells.