Dataset Information

The Impact of Heterogeneity on Single-Cell Sequencing.

ABSTRACT: The importance of diversity and cellular specialization is clear for many reasons, from population-level diversification, to improved resiliency to unforeseen stresses, to unique functions within metazoan organisms during development and differentiation. However, the level of cellular heterogeneity is just now becoming clear through the integration of genome-wide analyses and more cost effective Next Generation Sequencing (NGS). With easy access to single-cell NGS (scNGS), new opportunities exist to examine different levels of gene expression and somatic mutational heterogeneity, but these assays can generate yottabyte scale data. Here, we model the importance of heterogeneity for large-scale analysis of scNGS data, with a focus on the utilization in oncology and other diseases, providing a guide to aid in sample size and experimental design.

SUBMITTER: Goldman SL

PROVIDER: S-EPMC6405636 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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The Impact of Heterogeneity on Single-Cell Sequencing.

Goldman Samantha L SL MacKay Matthew M Afshinnekoo Ebrahim E Melnick Ari M AM Wu Shuxiu S Mason Christopher E CE

Frontiers in genetics 20190301

The importance of diversity and cellular specialization is clear for many reasons, from population-level diversification, to improved resiliency to unforeseen stresses, to unique functions within metazoan organisms during development and differentiation. However, the level of cellular heterogeneity is just now becoming clear through the integration of genome-wide analyses and more cost effective Next Generation Sequencing (NGS). With easy access to single-cell NGS (scNGS), new opportunities exis ...[more]

PMID: 30881372

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Project description:The current paradigm of osteoblast fate is that the majority undergo apoptosis, while some further differentiate into osteocytes and others flatten and cover bone surfaces as bone lining cells. Osteoblasts have been described to exhibit heterogeneous expression of a variety of osteoblast markers at both transcriptional and protein levels. To explore further this heterogeneity and its biological significance, Venus-positive (Venus+) cells expressing the fluorescent protein Venus under the control of the 2.3-kb Col1a1 promoter were isolated from newborn mouse calvariae and subjected to single-cell RNA sequencing. Functional annotation of the genes expressed in 272 Venus+ single cells indicated that Venus+ cells are osteoblasts that can be categorized into four clusters. Of these, three clusters (clusters 1 to 3) exhibited similarities in their expression of osteoblast markers, while one (cluster 4) was distinctly different. We identified a total of 1920 cluster-specific genes and pseudotime ordering analyses based on established concepts and known markers showed that clusters 1 to 3 captured osteoblasts at different maturational stages. Analysis of gene co-expression networks showed that genes involved in protein synthesis and protein trafficking between endoplasmic reticulum (ER) and Golgi are active in these clusters. However, the cells in these clusters were also defined by extensive heterogeneity of gene expression, independently of maturational stage. Cells of cluster 4 expressed Cd34 and Cxcl12 with relatively lower levels of osteoblast markers, suggesting that this cell type differs from actively bone-forming osteoblasts and retain or reacquire progenitor properties. Based on expression and machine learning analyses of the transcriptomes of individual osteoblasts, we also identified genes that may be useful as new markers of osteoblast maturational stages. Taken together, our data show much more extensive heterogeneity of osteoblasts than previously documented, with gene profiles supporting diversity of osteoblast functional activities and developmental fates. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

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Dataset Information

The Impact of Heterogeneity on Single-Cell Sequencing.

Publications

The Impact of Heterogeneity on Single-Cell Sequencing.

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