scholarly journals Lineage tracing on transcriptional landscapes links state to fate during differentiation

2018 ◽  
Author(s):  
Caleb Weinreb ◽  
Alejo Rodriguez-Fraticelli ◽  
Fernando Camargo ◽  
Allon M Klein
Keyword(s):  
Single Cell ◽  
Cell Biology ◽  
Cell Types ◽  
Lineage Tracing ◽  
Cell Potential ◽  
Monocyte Differentiation ◽  
Cell Assays ◽  
Transcriptional Landscape ◽  
Developmental Hierarchy

AbstractA challenge in stem cell biology is to associate molecular differences among progenitor cells with their capacity to generate mature cell types. Though the development of single cell assays allows for the capture of progenitor cell states in great detail, these assays cannot definitively link cell states to their long-term fate. Here, we use expressed DNA barcodes to clonally trace single cell transcriptomes dynamically during differentiation and apply this approach to the study of hematopoiesis. Our analysis identifies functional boundaries of cell potential early in the hematopoietic hierarchy and locates them on a continuous transcriptional landscape. We reconstruct a developmental hierarchy showing separate ontogenies for granulocytic subtypes and two routes to monocyte differentiation that leave a persistent imprint on mature cells. Finally, we use our approach to benchmark methods of dynamic inference from single-cell snapshots, and provide evidence of strong early fate biases dependent on cellular properties hidden from single-cell RNA sequencing.

scholarly journals Lineage tracing on transcriptional landscapes links state to fate during differentiation

Science ◽  
2020 ◽  
Vol 367 (6479) ◽  
pp. eaaw3381 ◽  
Author(s):  
Caleb Weinreb ◽  
Alejo Rodriguez-Fraticelli ◽  
Fernando D. Camargo ◽  
Allon M. Klein
Keyword(s):  
Single Cell ◽  
State Of The Art ◽  
Cell Types ◽  
Lineage Tracing ◽  
Dna Barcodes ◽  
Mature Cell ◽  
Monocyte Differentiation ◽  
Single Cell Rna Sequencing ◽  
Sister Cells ◽  
Transcriptional Landscape

A challenge in biology is to associate molecular differences among progenitor cells with their capacity to generate mature cell types. Here, we used expressed DNA barcodes to clonally trace transcriptomes over time and applied this to study fate determination in hematopoiesis. We identified states of primed fate potential and located them on a continuous transcriptional landscape. We identified two routes of monocyte differentiation that leave an imprint on mature cells. Analysis of sister cells also revealed cells to have intrinsic fate biases not detectable by single-cell RNA sequencing. Finally, we benchmarked computational methods of dynamic inference from single-cell snapshots, showing that fate choice occurs earlier than is detected by state-of the-art algorithms and that cells progress steadily through pseudotime with precise and consistent dynamics.

scholarly journals Short-read and long-read RNA sequencing of mouse hematopoietic stem cells at bulk and single-cell levels

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Xiuran Zheng ◽  
Dan Zhang ◽  
Mengying Xu ◽  
Wanqin Zeng ◽  
Ran Zhou ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Single Cell ◽  
Rna Sequencing ◽  
Cell Types ◽  
Lineage Tracing ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Long Read

AbstractHematopoietic stem cells (HSCs) lie at the top of the differentiation hierarchy. Although HSC and their immediate downstream, multipotent progenitors (MPP) have full multilineage differentiation capacity, only long-term (LT-) HSC has the capacity of long-term self-renewal. The heterogeneity within the HSC population is gradually acknowledged with the development of single-cell RNA sequencing and lineage tracing technologies. Transcriptional and post-transcriptional regulations play important roles in controlling the differentiation and self-renewal capacity within HSC population. Here we report a dataset comprising short- and long-read RNA sequencing for mouse long- and short-term HSC and MPP at bulk and single-cell levels. We demonstrate that integrating short- and long-read sequencing can facilitate the identification and quantification of known and unannotated isoforms. Thus, this dataset provides a groundwork for comprehensive and comparative studies on transcriptional diversity and heterogeneity within different HSC cell types.

scholarly journals Simultaneous trimodal single-cell measurement of transcripts, epitopes, and chromatin accessibility using TEA-seq

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Elliott Swanson ◽  
Cara Lord ◽  
Julian Reading ◽  
Alexander T Heubeck ◽  
Palak C Genge ◽  
...  
Keyword(s):  
Gene Regulation ◽  
Single Cell ◽  
Human Peripheral Blood ◽  
Single Cells ◽  
Cell Types ◽  
Chromatin Accessibility ◽  
Specific Gene ◽  
Test Case ◽  
Cell Assays ◽  
Paired Measurement

Single-cell measurements of cellular characteristics have been instrumental in understanding the heterogeneous pathways that drive differentiation, cellular responses to signals, and human disease. Recent advances have allowed paired capture of protein abundance and transcriptomic state, but a lack of epigenetic information in these assays has left a missing link to gene regulation. Using the heterogeneous mixture of cells in human peripheral blood as a test case, we developed a novel scATAC-seq workflow that increases signal-to-noise and allows paired measurement of cell surface markers and chromatin accessibility: integrated cellular indexing of chromatin landscape and epitopes, called ICICLE-seq. We extended this approach using a droplet-based multiomics platform to develop a trimodal assay that simultaneously measures transcriptomics (scRNA-seq), epitopes, and chromatin accessibility (scATAC-seq) from thousands of single cells, which we term TEA-seq. Together, these multimodal single-cell assays provide a novel toolkit to identify type-specific gene regulation and expression grounded in phenotypically defined cell types.

scholarly journals Cells expressing PAX8 are the main source of homeostatic regeneration of adult mouse endometrial epithelium and give rise to serous endometrial carcinoma

2020 ◽  
Vol 13 (10) ◽  
pp. dmm047035
Author(s):  
Dah-Jiun Fu ◽  
Andrea J. De Micheli ◽  
Mallikarjun Bidarimath ◽  
Lora H. Ellenson ◽  
Benjamin D. Cosgrove ◽  
...  
Keyword(s):  
Stem Cells ◽  
Epithelial Cells ◽  
Endometrial Carcinoma ◽  
Marrow Cell ◽  
Cell Types ◽  
Lineage Tracing ◽  
Cell Of Origin ◽  
Mouse Uterus ◽  
Endometrial Epithelium

ABSTRACTHumans and mice have cyclical regeneration of the endometrial epithelium. It is expected that such regeneration is ensured by tissue stem cells, but their location and hierarchy remain debatable. A number of recent studies have suggested the presence of stem cells in the mouse endometrial epithelium. At the same time, it has been reported that this tissue can be regenerated by stem cells of stromal/mesenchymal or bone marrow cell origin. Here, we describe a single-cell transcriptomic atlas of the main cell types of the mouse uterus and epithelial subset transcriptome and evaluate the contribution of epithelial cells expressing the transcription factor PAX8 to the homeostatic regeneration and malignant transformation of adult endometrial epithelium. According to lineage tracing, PAX8+ epithelial cells are responsible for long-term maintenance of both luminal and glandular epithelium. Furthermore, multicolor tracing shows that individual glands and contiguous areas of luminal epithelium are formed by clonal cell expansion. Inactivation of the tumor suppressor genes Trp53 and Rb1 in PAX8+ cells, but not in FOXJ1+ cells, leads to the formation of neoplasms with features of serous endometrial carcinoma, one of the most aggressive types of human endometrial malignancies. Taken together, our results show that the progeny of single PAX8+ cells represents the main source of regeneration of the adult endometrial epithelium. They also provide direct experimental genetic evidence for the key roles of the P53 and RB pathways in the pathogenesis of serous endometrial carcinoma and suggest that PAX8+ cells represent the cell of origin of this neoplasm.

Cellular phenotypes of normal and leukemic hemopoietic cells determined by analysis with selected antibody combinations

Blood ◽  
1980 ◽  
Vol 56 (3) ◽  
pp. 430-441 ◽  
Author(s):  
G Janossy ◽  
FJ Bollum ◽  
KF Bradstock ◽  
J Ashley
Keyword(s):  
Bone Marrow ◽  
Monoclonal Antibodies ◽  
Single Cell ◽  
Normal Cell ◽  
Cell Types ◽  
Precursor Cells ◽  
Leukemic Cells ◽  
Double Staining ◽  
Cell Assays ◽  
Cellular Phenotypes

Abstract Individual leukemic cells and the corresponding rare normal cell types in nonleukemic bone marrow were analyzed with various combinations of antisera (labeled with different fluorochromes: TRITC and FITC). Double staining for membrane Ia-like molecules (TRITC) and nuclear terminal transferase (FITC) was a very useful combination that distinguished common non-T, non-B ALL (Ia+,TdT+) and thymic ALL (Ia-,TdT+) from the rare cases of B ALL (Ia+,TdT-) and from AML (frequently Ia+, TdT-; in some cases Ia-, TdT-). Additional antisera (such as anti-ALL, anti- HuTLA, anti-immunoglobulin reagents, etc.) confirmed the diagnosis and further characterized the leukemic blasts. Ia+,TdT+ cells could be observed in low numbers in normal and nonleukemic regenerating marrow and were probably normal precursor cells; this reagent combinations was, therefore, not useful for monitoring residual non-T, non-B ALL blasts in treated patients. Other marker combinations detecting pre-B ALL blasts (double staining for cytoplasmic IgM and nuclear TdT) and Thy-ALL blasts (HuTLA+,TdT+) were, however, virtually leukemia specific in the bone marrow and could be used to effectively monitor residual leukemic cells throughout the disease. These combined single-cell assays are not only economical and informative but are also important for assessing the heterogeneity of leukemia and for standardizing new mouse or rat monoclonal antibodies for diagnosis.

scholarly journals Kinetics of adult hematopoietic stem cell differentiation in vivo

2018 ◽  
Vol 215 (11) ◽  
pp. 2815-2832 ◽  
Author(s):  
Samik Upadhaya ◽  
Catherine M. Sawai ◽  
Efthymia Papalexi ◽  
Ali Rashidfarrokhi ◽  
Geunhyo Jang ◽  
...  
Keyword(s):  
Stem Cell Differentiation ◽  
Cell Types ◽  
Lineage Tracing ◽  
Intermediate Cell ◽  
Hematopoietic Stem ◽  
Hematopoietic Stem Cell Differentiation ◽  
Kinetics Of ◽  
Kinetics In Vivo

Adult hematopoiesis has been studied in terms of progenitor differentiation potentials, whereas its kinetics in vivo is poorly understood. We combined inducible lineage tracing of endogenous adult hematopoietic stem cells (HSCs) with flow cytometry and single-cell RNA sequencing to characterize early steps of hematopoietic differentiation in the steady-state. Labeled cells, comprising primarily long-term HSCs and some short-term HSCs, produced megakaryocytic lineage progeny within 1 wk in a process that required only two to three cell divisions. Erythroid and myeloid progeny emerged simultaneously by 2 wk and included a progenitor population with expression features of both lineages. Myeloid progenitors at this stage showed diversification into granulocytic, monocytic, and dendritic cell types, and rare intermediate cell states could be detected. In contrast, lymphoid differentiation was virtually absent within the first 3 wk of tracing. These results show that continuous differentiation of HSCs rapidly produces major hematopoietic lineages and cell types and reveal fundamental kinetic differences between megakaryocytic, erythroid, myeloid, and lymphoid differentiation.

scholarly journals Applications of Single-Cell Omics to Dissect Tumor Microenvironment

2020 ◽  
Vol 11 ◽  
Author(s):  
Tingting Guo ◽  
Weimin Li ◽  
Xuyu Cai
Keyword(s):  
Tumor Microenvironment ◽  
Single Cell ◽  
Immune Cells ◽  
Immune Cell ◽  
Cell Types ◽  
Immune Checkpoint Blockade ◽  
Lineage Tracing ◽  
Great Success ◽  
Novel Technologies ◽  
Single Cell Sequencing

The recent technical and computational advances in single-cell sequencing technologies have significantly broaden our toolkit to study tumor microenvironment (TME) directly from human specimens. The TME is the complex and dynamic ecosystem composed of multiple cell types, including tumor cells, immune cells, stromal cells, endothelial cells, and other non-cellular components such as the extracellular matrix and secreted signaling molecules. The great success on immune checkpoint blockade therapy has highlighted the importance of TME on anti-tumor immunity and has made it a prime target for further immunotherapy strategies. Applications of single-cell transcriptomics on studying TME has yielded unprecedented resolution of the cellular and molecular complexity of the TME, accelerating our understanding of the heterogeneity, plasticity, and complex cross-interaction between different cell types within the TME. In this review, we discuss the recent advances by single-cell sequencing on understanding the diversity of TME and its functional impact on tumor progression and immunotherapy response driven by single-cell sequencing. We primarily focus on the major immune cell types infiltrated in the human TME, including T cells, dendritic cells, and macrophages. We further discuss the limitations of the existing methodologies and the prospects on future studies utilizing single-cell multi-omics technologies. Since immune cells undergo continuous activation and differentiation within the TME in response to various environmental cues, we highlight the importance of integrating multimodal datasets to enable retrospective lineage tracing and epigenetic profiling of the tumor infiltrating immune cells. These novel technologies enable better characterization of the developmental lineages and differentiation states that are critical for the understanding of the underlying mechanisms driving the functional diversity of immune cells within the TME. We envision that with the continued accumulation of single-cell omics datasets, single-cell sequencing will become an indispensable aspect of the immune-oncology experimental toolkit. It will continue to drive the scientific innovations in precision immunotherapy and will be ultimately adopted by routine clinical practice in the foreseeable future.

scholarly journals Mouse telomerase reverse transcriptase (mTert) expression marks slowly cycling intestinal stem cells

2010 ◽  
Vol 108 (1) ◽  
pp. 179-184 ◽  
Author(s):  
Robert K. Montgomery ◽  
Diana L. Carlone ◽  
Camilla A. Richmond ◽  
Loredana Farilla ◽  
Mariette E. G. Kranendonk ◽  
...  
Keyword(s):  
Stem Cells ◽  
Reverse Transcriptase ◽  
Tissue Injury ◽  
Cell Types ◽  
Lineage Tracing ◽  
Intestinal Stem Cells ◽  
Intestinal Cell ◽  
Stem Cell Population ◽  
Telomerase Reverse Transcriptase

The intestinal epithelium is maintained by a population of rapidly cycling (Lgr5+) intestinal stem cells (ISCs). It has been postulated, however, that slowly cycling ISCs must also be present in the intestine to protect the genome from accumulating deleterious mutations and to allow for a response to tissue injury. Here, we identify a subpopulation of slowly cycling ISCs marked by mouse telomerase reverse transcriptase (mTert) expression that can give rise to Lgr5+ cells. mTert-expressing cells distribute in a pattern along the crypt–villus axis similar to long-term label-retaining cells (LRCs) and are resistant to tissue injury. Lineage-tracing studies demonstrate that mTert+ cells give rise to all differentiated intestinal cell types, persist long term, and contribute to the regenerative response following injury. Consistent with other highly regenerative tissues, our results demonstrate that a slowly cycling stem cell population exists within the intestine.

scholarly journals TEA-seq: a trimodal assay for integrated single cell measurement of transcription, epitopes, and chromatin accessibility

2020 ◽  
Author(s):  
Elliott Swanson ◽  
Cara Lord ◽  
Julian Reading ◽  
Alexander T. Heubeck ◽  
Adam K. Savage ◽  
...  
Keyword(s):  
Cell Surface ◽  
Single Cell ◽  
Human Peripheral Blood ◽  
Signal To Noise Ratio ◽  
Single Cells ◽  
Cell Types ◽  
Chromatin Accessibility ◽  
Specific Gene ◽  
Test Case ◽  
Cell Assays

AbstractSingle-cell measurements of cellular characteristics have been instrumental in understanding the heterogeneous pathways that drive differentiation, cellular responses to extracellular signals, and human disease states. scATAC-seq has been particularly challenging due to the large size of the human genome and processing artefacts resulting from DNA damage that are an inherent source of background signal. Downstream analysis and integration of scATAC-seq with other single-cell assays is complicated by the lack of clear phenotypic information linking chromatin state and cell type. Using the heterogeneous mixture of cells in human peripheral blood as a test case, we developed a novel scATAC-seq workflow that increases the signal-to-noise ratio and allows simultaneous measurement of cell surface markers: Integrated Cellular Indexing of Chromatin Landscape and Epitopes (ICICLE-seq). We extended this approach using a droplet-based multiomics platform to develop a trimodal assay to simultaneously measure Transcriptomic state (scRNA-seq), cell surface Epitopes, and chromatin Accessibility (scATAC-seq) from thousands of single cells, which we term TEA-seq. Together, these multimodal single-cell assays provide a novel toolkit to identify type-specific gene regulation and expression grounded in phenotypically defined cell types.

scholarly journals A Single-Cell Atlas and Lineage Analysis of the Adult Drosophila Ovary

2019 ◽  
Author(s):  
Katja Rust ◽  
Lauren Byrnes ◽  
Kevin Shengyang Yu ◽  
Jason S. Park ◽  
Julie B. Sneddon ◽  
...  
Keyword(s):  
Stem Cells ◽  
Single Cell ◽  
Cell Types ◽  
Somatic Tissue ◽  
Lineage Tracing ◽  
Major Cell Type ◽  
Follicle Stem Cells ◽  
Drosophila Ovary ◽  
Adult Drosophila ◽  
Lineage Analysis

AbstractThe Drosophila ovary is a widely used model for germ cell and somatic tissue biology. We have used single-cell RNA-sequencing to build a comprehensive cell atlas of the adult Drosophila ovary containing unique transcriptional profiles for every major cell type in the ovary, including the germline and follicle stem cells. Using this atlas we identify novel tools for identification and manipulation of known and novel cell types and perform lineage tracing to test cellular relationships of previously unknown cell types. By this we discovered a new form of cellular plasticity in which inner germarial sheath cells convert to follicle stem cells in response to starvation.Graphical Abstract

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