scholarly journals Single-cell perturbation dissects transcription factor control of progression speed and trajectory choice in early T-cell development

2021 ◽  
Author(s):  
Wen Zhou ◽  
Fan Gao ◽  
Maile Romero-Wolf ◽  
Suin Jo ◽  
Ellen V. Rothenberg
Keyword(s):  
T Cell ◽  
Single Cell ◽  
Single Cells ◽  
T Cell Development ◽  
Cell Development ◽  
Lineage Commitment ◽  
Functional Roles ◽  
Two Stages ◽  
Developmental Block

In early T-cell development, single cells dynamically shift expression of multiple transcription factors (TFs) during transition from multipotentiality to T-lineage commitment, but the functional roles of many TFs have been obscure. Here, synchronized in vitro differentiation systems, scRNA-seq with batch indexing, and controlled gene-disruption strategies have unraveled single-cell impacts of perturbing individual TFs at two stages in early T-cell development. Single-cell CRISPR perturbation revealed that early-acting TFs Bcl11a, Erg, Spi1 (PU.1), Gata3, and Tcf7 (TCF1) each play individualized roles promoting or retarding T-lineage progression and suppressing alternative trajectories, collectively determining population dynamics and path topologies. Later, during T-lineage commitment, cells prevented from expressing TF Bcl11b "realized" this abnormality not with a developmental block, but by shifting into a divergent path via bZIP and Sox TF activation as well as E protein antagonism, finally exiting the T-lineage trajectory. These TFs thus exert a network of impacts to control progression kinetics, trajectories, and differentiation outcomes of early pro-T cells.

scholarly journals Hierarchy of Notch–Delta interactions promoting T cell lineage commitment and maturation

2007 ◽  
Vol 204 (2) ◽  
pp. 331-343 ◽  
Author(s):  
Valerie Besseyrias ◽  
Emma Fiorini ◽  
Lothar J. Strobl ◽  
Ursula Zimber-Strobl ◽  
Alexis Dumortier ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Development ◽  
Cell Lineage ◽  
Cell Development ◽  
Lineage Commitment ◽  
Binding Studies ◽  
Maturation In Vitro ◽  
Cell Commitment

Notch1 (N1) receptor signaling is essential and sufficient for T cell development, and recently developed in vitro culture systems point to members of the Delta family as being the physiological N1 ligands. We explored the ability of Delta1 (DL1) and DL4 to induce T cell lineage commitment and/or maturation in vitro and in vivo from bone marrow (BM) precursors conditionally gene targeted for N1 and/or N2. In vitro DL1 can trigger T cell lineage commitment via either N1 or N2. N1- or N2-mediated T cell lineage commitment can also occur in the spleen after short-term BM transplantation. However, N2–DL1–mediated signaling does not allow further T cell maturation beyond the CD25+ stage due to a lack of T cell receptor β expression. In contrast to DL1, DL4 induces and supports T cell commitment and maturation in vitro and in vivo exclusively via specific interaction with N1. Moreover, comparative binding studies show preferential interaction of DL4 with N1, whereas binding of DL1 to N1 is weak. Interestingly, preferential N1–DL4 binding reflects reduced dependence of this interaction on Lunatic fringe, a glycosyl transferase that generally enhances the avidity of Notch receptors for Delta ligands. Collectively, our results establish a hierarchy of Notch–Delta interactions in which N1–DL4 exhibits the greatest capacity to induce and support T cell development.

scholarly journals Single-Cell Analysis Reveals Regulatory Gene Expression Dynamics Leading to Lineage Commitment in Early T Cell Development

Cell Systems ◽  
2019 ◽  
Vol 9 (4) ◽  
pp. 321-337.e9 ◽  
Author(s):  
Wen Zhou ◽  
Mary A. Yui ◽  
Brian A. Williams ◽  
Jina Yun ◽  
Barbara J. Wold ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cell ◽  
Single Cell ◽  
Single Cell Analysis ◽  
Regulatory Gene ◽  
T Cell Development ◽  
Cell Development ◽  
Lineage Commitment ◽  
Cell Analysis ◽  
Gene Expression Dynamics

scholarly journals Delta-like 4 is the essential, nonredundant ligand for Notch1 during thymic T cell lineage commitment

2008 ◽  
Vol 205 (11) ◽  
pp. 2515-2523 ◽  
Author(s):  
Ute Koch ◽  
Emma Fiorini ◽  
Rui Benedito ◽  
Valerie Besseyrias ◽  
Karin Schuster-Gossler ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
T Cell Development ◽  
Cell Lineage ◽  
Cell Development ◽  
Lineage Commitment ◽  
Thymic Epithelial Cells ◽  
Hematopoietic Progenitors ◽  
Immature B Cells

Thymic T cell lineage commitment is dependent on Notch1 (N1) receptor–mediated signaling. Although the physiological ligands that interact with N1 expressed on thymic precursors are currently unknown, in vitro culture systems point to Delta-like 1 (DL1) and DL4 as prime candidates. Using DL1- and DL4-lacZ reporter knock-in mice and novel monoclonal antibodies to DL1 and DL4, we show that DL4 is expressed on thymic epithelial cells (TECs), whereas DL1 is not detected. The function of DL4 was further explored in vivo by generating mice in which DL4 could be specifically inactivated in TECs or in hematopoietic progenitors. Although loss of DL4 in hematopoietic progenitors did not perturb thymus development, inactivation of DL4 in TECs led to a complete block in T cell development coupled with the ectopic appearance of immature B cells in the thymus. These immature B cells were phenotypically indistinguishable from those developing in the thymus of conditional N1 mutant mice. Collectively, our results demonstrate that DL4 is the essential and nonredundant N1 ligand responsible for T cell lineage commitment. Moreover, they strongly suggest that N1-expressing thymic progenitors interact with DL4-expressing TECs to suppress B lineage potential and to induce the first steps of intrathymic T cell development.

3040 – IN VITRO RECAPITULATION OF MURINE T CELL DEVELOPMENT FROM SINGLE HEMATOPOIETIC STEM CELLS

2020 ◽  
Vol 88 ◽  
pp. S51
Author(s):  
Victoria Sun ◽  
Amelie Montel-Hagen ◽  
David Casero ◽  
Steven Tsai ◽  
Alexandre Zampieri ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cell ◽  
Hematopoietic Stem Cells ◽  
T Cell Development ◽  
Cell Development ◽  
Hematopoietic Stem

Single-Cell Transcriptomics Reveals Discrete Steps in Regulatory T Cell Development in the Human Thymus

2021 ◽  
pp. ji2100506
Author(s):  
Florencia Morgana ◽  
Rianne Opstelten ◽  
Manon C. Slot ◽  
Andrew M. Scott ◽  
René A. W. van Lier ◽  
...  
Keyword(s):  
T Cell ◽  
Single Cell ◽  
Regulatory T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Human Thymus

scholarly journals Runx1 and Runx3 drive progenitor to T-lineage transcriptome conversion in mouse T cell commitment via dynamic genomic site switching

2021 ◽  
Vol 118 (4) ◽  
pp. e2019655118 ◽  
Author(s):  
Boyoung Shin ◽  
Hiroyuki Hosokawa ◽  
Maile Romero-Wolf ◽  
Wen Zhou ◽  
Kaori Masuhara ◽  
...  
Keyword(s):  
T Cell ◽  
Target Genes ◽  
Single Cells ◽  
T Cell Development ◽  
Cell Development ◽  
Dominant Factor ◽  
Runt Domain ◽  
Binding Behavior ◽  
Regulation Function ◽  
Cell Commitment

Runt domain-related (Runx) transcription factors are essential for early T cell development in mice from uncommitted to committed stages. Single and double Runx knockouts via Cas9 show that target genes responding to Runx activity are not solely controlled by the dominant factor, Runx1. Instead, Runx1 and Runx3 are coexpressed in single cells; bind to highly overlapping genomic sites; and have redundant, collaborative functions regulating genes pivotal for T cell development. Despite stable combined expression levels across pro-T cell development, Runx1 and Runx3 preferentially activate and repress genes that change expression dynamically during lineage commitment, mostly activating T-lineage genes and repressing multipotent progenitor genes. Furthermore, most Runx target genes are sensitive to Runx perturbation only at one stage and often respond to Runx more for expression transitions than for maintenance. Contributing to this highly stage-dependent gene regulation function, Runx1 and Runx3 extensively shift their binding sites during commitment. Functionally distinct Runx occupancy sites associated with stage-specific activation or repression are also distinguished by different patterns of partner factor cobinding. Finally, Runx occupancies change coordinately at numerous clustered sites around positively or negatively regulated targets during commitment. This multisite binding behavior may contribute to a developmental “ratchet” mechanism making commitment irreversible.

scholarly journals Dynamic regulation of chromatin organizer SATB1 via TCR-induced alternative promoter switch during T-cell development

2020 ◽  
Vol 48 (11) ◽  
pp. 5873-5890
Author(s):  
Indumathi Patta ◽  
Ayush Madhok ◽  
Satyajeet Khare ◽  
Kamalvishnu P Gottimukkala ◽  
Anjali Verma ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Alternative Promoter ◽  
In Vitro Translation ◽  
Alternative Promoters ◽  
Transcript Variants ◽  
P2 Promoter ◽  
Specific Regulation

Abstract The chromatin organizer SATB1 is highly enriched in thymocytes and is essential for T-cell development. Although SATB1 regulates a large number of genes important for T-cell development, the mechanism(s) regulating expression of SATB1 during this process remain elusive. Using chromatin immune precipitation-seq-based occupancy profiles of H3K4me3 and H3Kme1 at Satb1 gene locus, we predicted four different alternative promoters of Satb1 in mouse thymocytes and characterized them. The expression of Satb1 transcript variants with distinct 5′ UTRs occurs in a stage-specific manner during T-cell development and is dependent on TCR signaling. The observed discrepancy between the expression levels of SATB1 mRNA and protein in developing thymocytes can be explained by the differential translatability of Satb1 transcript variants as confirmed by polysome profiling and in vitro translation assay. We show that Satb1 alternative promoters exhibit lineage-specific chromatin accessibility during T-cell development from progenitors. Furthermore, TCF1 regulates the Satb1 P2 promoter switch during CD4SP development, via direct binding to the Satb1 P2 promoter. CD4SP T cells from TCF1 KO mice exhibit downregulation of P2 transcript variant expression as well as low levels of SATB1 protein. Collectively, these results provide unequivocal evidence toward alternative promoter switch-mediated developmental stage-specific regulation of SATB1 in thymocytes.

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