Single-cell transcriptomics reveals a new dynamical function of transcription factors during embryonic hematopoiesis

Bergiers, Isabelle; Andrews, Tallulah; Bölükbaşı, Özge Vargel; Buneß, Andreas; Janosz, Ewa; López-Anguita, Natalia; Ganter, Kerstin; Kosim, Kinga; Celen, Cemre; Perçin, Gülce Itır; Collier, Paul; Baying, Bianka; Beneš, Vladimı́r; Hemberg, Martin; Lancrin, Christophe

doi:10.7554/elife.29312

Cited by 41 publications

(84 citation statements)

References 64 publications

(91 reference statements)

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“…This experiment was performed exactly as described previously 69 using the Fluidigm Biomark system. Data analysis was also done as before 69 .…”

Section: Single-cell Q-rt-pcr Analysismentioning

confidence: 99%

“…To generate the iStab2 ESC line, A2loxEmpty ES cells 69 Twenty-four hours after the transfection, cells were treated with 200μg/ml HygromycinB (CalBiochem, #400051) in the morning and evening for two days and then only in the morning for 5 more days. After 7 days of antibiotic treatment, cells were FACS Aria sorted for BFP on 96-well plate of mouse embryonic fibroblasts.…”

Section: Generation Of Esc Lines and In Vitro Esc Differentiation Intmentioning

confidence: 99%

“…ES cells were then subjected to in vitro differentiation into Embryoid bodies to produce blood, endothelial and vascular smooth muscle cells following the same protocol described previously 69 .…”

Section: Generation Of Esc Lines and In Vitro Esc Differentiation Intmentioning

confidence: 99%

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Single-cell atlas of major haematopoietic tissues sheds light on blood cell formation from embryonic endothelium

Shvartsman

Pavlovich

Oatley

et al. 2019

Preprint

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The Yolk Sac (YS) and Aorta-Gonad-Mesonephros (AGM) are two major haematopoietic regions during embryonic development. Interestingly, AGM is the only one generating haematopoietic stem cells (HSCs). To identify the difference between AGM and YS, we compared them using singlecell RNA sequencing between 9.5 and 11.5 days of mouse embryonic development and identified cell populations using CONCLUS, a new computational tool. The AGM was the only one containing neurons and a specific mesenchymal population, while the YS major component was an epithelial population expressing liver marker genes. In addition, the YS contained a major endothelial population expressing Stab2, a hyaluronan receptor, also highly expressed by liver endothelium. We demonstrated that the YS haematopoietic potential was restricted to Stab2negative cells and that ectopic expression of Stab2 could reduce blood cell formation from endothelium. Our results indicate that the AGM is a tissue more favourable to HSCs development than the YS because of its microenvironment and the nature of its endothelial cells.

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“…This experiment was performed exactly as described previously 69 using the Fluidigm Biomark system. Data analysis was also done as before 69 .…”

Section: Single-cell Q-rt-pcr Analysismentioning

confidence: 99%

Section: Generation Of Esc Lines and In Vitro Esc Differentiation Intmentioning

confidence: 99%

See 1 more Smart Citation

Single-cell atlas of major haematopoietic tissues sheds light on blood cell formation from embryonic endothelium

Shvartsman

Pavlovich

Oatley

et al. 2019

Preprint

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“…The widely used method is to quantify endothelial and hematopoietic marker expressing cells. This can be done by gene expression analysis at the single-cell level (Bergiers et al ., 2018). Also, protein expression change can be used to assess EHT progression by flow cytometry analysis of endothelial markers such as VE-Cadherin, CD31 and hematopoietic markers such as CD41, CD45, CD43.…”

Section: Introductionmentioning

confidence: 99%

“…Here, by combining in vitro time-lapse imaging of an adherent BL-CFC culture with automatic image analysis we introduce a simple and efficient method to quantify those round cells during a culture period, which gives a direct measure of the number of cells undergoing EHT. This protocol enables us to easily test novel parameters affecting EHT rate such as over-expression of certain transcription factors (Bergiers et al ., 2018) or testing pathway inhibiting small molecules in the culture media (Vargel et al ., 2016). Below, we describe the details of the time-lapse microscopy of BL-CFC culture and image analysis to assess the number of cells underwent EHT.…”

Section: Introductionmentioning

confidence: 99%

Quantification of Mouse Hematopoietic Progenitors’ Formation Using Time-lapse Microscopy and Image Analysis

et al. 2019

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In vitro differentiation of mouse embryonic stem cells (mESCs) towards blood cells constitutes a well-established system to study the endothelial-to-hematopoietic transition (EHT) at the onset of blood development. Assessing the emergence of small non-adherent round blood cells in the culture without disturbing it is essential to evaluate the progression of EHT and also to test conditions potentially enhancing or repressing this process. Here, we describe how to quantify the formation of mouse hematopoietic progenitors during EHT in normal conditions or following over-expression of eight essential transcription factors using time-lapse microscopy and image analysis.

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Combined Analysis of Transcriptome and T-Cell Receptor Alpha and Beta (TRA/TRB) Repertoire in Paucicellular Samples at the Single-Cell Level

Litjens

Langerak

Azmani

et al. 2022

Methods in Molecular Biology

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With the advent of next-generation sequencing (NGS) methodologies, the total repertoires of B and T cells can be disclosed in much more detail than ever before. Even though many of these strategies do provide in-depth and high-resolution information of the immunoglobulin (IG) and/or T-cell receptor (TR) repertoire, one clear disadvantage is that the IG/TR profiles cannot be connected to individual cells. Single-cell technologies do allow to study the IG/TR repertoire at the individual cell level. This is especially relevant in cell samples in which much heterogeneity of the cell population is expected. By combining the IG/TR repertoire with transcriptome data, the reactivity of the B or T cell can be associated with activation or maturation stages. An additional advantage of such single-cell technologies is that the combination of both IG and both TR chains can be studied on a per cell basis, which better reflects the antigen receptor reactivity of cells. Here we present the ICELL8 single-cell method for the parallel analysis of the TR repertoire and transcriptome, which is especially useful in samples that contain relatively few cells.

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Single-cell transcriptomics reveals a new dynamical function of transcription factors during embryonic hematopoiesis

Cited by 41 publications

References 64 publications

Single-cell atlas of major haematopoietic tissues sheds light on blood cell formation from embryonic endothelium

Single-cell atlas of major haematopoietic tissues sheds light on blood cell formation from embryonic endothelium

Quantification of Mouse Hematopoietic Progenitors’ Formation Using Time-lapse Microscopy and Image Analysis

Combined Analysis of Transcriptome and T-Cell Receptor Alpha and Beta (TRA/TRB) Repertoire in Paucicellular Samples at the Single-Cell Level

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