Modulation of microRNA expression in human T-cell development: targeting of NOTCH3 by miR-150

Blood ◽  
2011 ◽  
Vol 117 (26) ◽  
pp. 7053-7062 ◽  
Author(s):  
Margherita Ghisi ◽  
Alberto Corradin ◽  
Katia Basso ◽  
Chiara Frasson ◽  
Valentina Serafin ◽  
...  
Keyword(s):  
T Cell ◽  
Expression Profiles ◽  
T Cell Development ◽  
Notch Pathway ◽  
Cell Development ◽  
Microrna Expression ◽  
Double Positive ◽  
Molecular Control ◽  
Human T Cell ◽  
Single Positive

Abstract Ontogenesis of T cells in the thymus is a complex process whose molecular control is poorly understood. The present study investigated microRNAs involved in human thymocyte differentiation by comparing the microRNA expression profiles of thymocytes at the double-positive, single-positive CD4+ and single-positive CD8+ maturation stages. Microarray analysis showed that each thymocyte population displays a distinct microRNA expression profile that reflects their developmental relationships. Moreover, analysis of small-RNA libraries generated from human unsorted and double-positive thymocytes and from mature peripheral CD4+ and CD8+ T lymphocytes, together with the microarray data, indicated a trend toward up-regulation of microRNA expression during T-cell maturation after the double-positive stage and revealed a group of microRNAs regulated during normal T-cell development, including miR-150, which is strongly up-regulated as maturation progresses. We showed that miR-150 targets NOTCH3, a member of the Notch receptor family that plays important roles both in T-cell differentiation and leukemogenesis. Forced expression of miR-150 reduces NOTCH3 levels in T-cell lines and has adverse effects on their proliferation and survival. Overall, these findings suggest that control of the Notch pathway through miR-150 may have an important impact on T-cell development and physiology.

Altered thymocyte and T cell development in neonatal mice with hyperoxia-induced lung injury

2018 ◽  
Vol 46 (4) ◽  
pp. 441-449
Author(s):  
Sowmya Angusamy ◽  
Tamer Mansour ◽  
Mohammed Abdulmageed ◽  
Rachel Han ◽  
Brian C. Schutte ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Lung Injury ◽  
T Cell Development ◽  
Adaptive Immune System ◽  
Cell Development ◽  
Thymocyte Development ◽  
Double Positive ◽  
Neonatal Mice ◽  
Single Positive

Abstract Background: The adaptive immune system of neonates is relatively underdeveloped. The thymus is an essential organ for adaptive T cell development and might be affected during the natural course of oxygen induced lung injury. The effect of prolonged hyperoxia on the thymus, thymocyte and T cell development, and its proliferation has not been studied extensively. Methods: Neonatal mice were exposed to 85% oxygen (hyperoxia) or room air (normoxia) up to 28 days. Flow cytometry using surface markers were used to assay for thymocyte development and proliferation. Results: Mice exposed to prolonged hyperoxia had evidence of lung injury associated alveolar simplification, a significantly lower mean weight, smaller thymic size, lower mean thymocyte count and higher percentage of apoptotic thymocytes. T cells subpopulation in the thymus showed a significant reduction in the count and proliferation of double positive and double negative T cells. There was a significant reduction in the count and proliferation of single positive CD4+ and CD8+ T cells. Conclusions: Prolonged hyperoxia in neonatal mice adversely affected thymic size, thymocyte count and altered the distribution of T cells sub-populations. These results are consistent with the hypothesis that prolonged hyperoxia causes defective development of T cells in the thymus.

scholarly journals VßGene Family Usage in Spontaneous Lymphomas of AKR Mice: Evidence for Defective Clonal Deletion

1992 ◽  
Vol 2 (2) ◽  
pp. 95-101 ◽  
Author(s):  
Cees de Heer ◽  
Bernard de Geus ◽  
Henk-Jan Schuurma ◽  
Henk Van Loveren ◽  
Jan Rozing
Keyword(s):  
T Cell ◽  
T Cell Development ◽  
Cell Receptor ◽  
Cell Development ◽  
Mesenteric Lymph Nodes ◽  
Double Positive ◽  
Clonal Deletion ◽  
Lymphoma Cells ◽  
Single Positive ◽  
Akr Mice

T-cell receptor (TCR)ß-chain usage and expression of the CD3, CD4, and CD8 differentiation antigens were analyzed in 14 spontaneous AKR lymphomas. Lymphoma cells massively infiltrated and/or proliferated in the organs analyzed (thymus, spleen, and mesenteric lymph nodes), giving rise to a loss of organ structure. One lymphoma occurred only in the thymus, and failed to express CD3, CD4, and CD8. All other lymphomas expressed the CD3/TCR complex. With respect to CD4 and CD8 expression, the lymphomas were either double-negative (DN), double-positive (DP), or single-positive (SP). The frequency of DP (CD4+8+) lymphomas was low compared to the frequency of DP thymocytes in a normal AKR thymus. A substantial heterogeneity was seen in the intensity of CD4 and CD8 expression among various lymphomas, which was independent of the level of CD3 expression. Considering TCR Vßgene family usage, 2 out of 14 lymphomas expressed Vß6. Normally, Vß6+thymocytes are deleted from the thymocyte pool at the immature DP stage of T-cell development in AKR mice. These data support the hypothesis that the lymphocytes in the immature DP stage of T-cell development are susceptible to the induction of AKR lymphomagenesis. The presence of Vß6+lymphoma cells indicates that the lymphomagenesis is accompanied by a defective clonal deletion of cells expressing a possible autoreactive TCR.

scholarly journals Disregulated expression of the transcription factor ThPOK during T-cell development leads to high incidence of T-cell lymphomas

2015 ◽  
Vol 112 (25) ◽  
pp. 7773-7778 ◽  
Author(s):  
Hyung-Ok Lee ◽  
Xiao He ◽  
Jayati Mookerjee-Basu ◽  
Dai Zhongping ◽  
Xiang Hua ◽  
...  
Keyword(s):  
Transcription Factor ◽  
T Cell ◽  
Lymphoblastic Leukemia ◽  
T Cell Development ◽  
Cell Receptor ◽  
Cell Development ◽  
Thymocyte Development ◽  
Tcr Signaling ◽  
Double Positive ◽  
Human T Cell

The transcription factor T-helper-inducing POZ/Krueppel-like factor (ThPOK, encoded by the Zbtb7b gene) plays widespread and critical roles in T-cell development, particularly as the master regulator of CD4 commitment. Here we show that mice expressing a constitutive T-cell–specific ThPOK transgene (ThPOKconst mice) develop thymic lymphomas. These tumors resemble human T-cell acute lymphoblastic leukemia (T-ALL), in that they predominantly exhibit activating Notch1 mutations. Lymphomagenesis is prevented if thymocyte development is arrested at the DN3 stage by recombination-activating gene (RAG) deficiency, but restored by introduction of a T-cell receptor (TCR) transgene or by a single injection of anti-αβTCR antibody into ThPOKconst RAG-deficient mice, which promotes development to the CD4+8+ (DP) stage. Hence, TCR signals and/or traversal of the DN (double negative) > DP (double positive) checkpoint are required for ThPOK-mediated lymphomagenesis. These results demonstrate a novel link between ThPOK, TCR signaling, and lymphomagenesis. Finally, we present evidence that ectopic ThPOK expression gives rise to a preleukemic and self-perpetuating DN4 lymphoma precursor population. Our results collectively define a novel role for ThPOK as an oncogene and precisely map the stage in thymopoiesis susceptible to ThPOK-dependent tumor initiation.

Heme Metabolic Gene Expression In FLVCR-Knockout Mice During T Cell Development

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2787-2787
Author(s):  
Mary Philip ◽  
Alexandra R. Zaballa ◽  
Blake T. Hovde ◽  
Janis L. Abkowitz
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Transcriptional Level ◽  
Mrna Levels ◽  
Double Positive ◽  
Metabolic Gene Expression ◽  
Free Heme ◽  
Single Positive

Abstract Abstract 2787 Heme is essential for nearly every organism and cell. However, free heme can induce free radical formation and cellular damage, therefore cells must carefully regulate heme levels. The feline leukemia virus subgroup C receptor (FLVCR) exports heme from cells. Conditional deletion of Flvcr has been shown to cause progressive anemia in neonatal and adult mice (Science 319:825-8, 2008). Recently, we developed a transplant model in which developing lymphocytes lacked FLVCR while erythroid cells expressed FLVCR, preventing anemia, and found that CD4 and CD8 peripheral T cells were severely decreased while B cell numbers were normal. We further demonstrated that FLVCR-knockout thymocytes were blocked at the CD4CD8 double-positive (DP) stage (Blood [ASH Annual Meeting Abstracts] 114: 913, 2009). We hypothesized that developing T cells lacking FLVCR are arrested at the DP stage because of increased intracellular free heme (IFH). While heme is required for erythroid function, little is known about the role of heme in T cell development. Real-time dynamic quantification of IFH in vivo or from ex vivo tissue is a major challenge in heme biology. We reasoned that by measuring the expression of genes transcriptionally-regulated by heme, we could indirectly assess IFH. Three proteins are key regulators of IFH in non-erythroid cells: aminolevulinic acid synthase-1 (ALAS1) is the rate-limiting enzyme in heme synthesis, FLVCR exports heme, and heme oxygenase-1 (HMOX1) degrades heme. Normal thymic T cell development proceeds from the CD4CD8 double-negative (DN) to the CD4CD8 double-positive (DP) stage, which then go on to either the CD4 single-positive (CD4SP) or CD8 single-positive (CD8SP) stage. We flow-sorted cells from each stage and used multiplex quantitative PCR (qPCR) to determine that all three genes were expressed at higher levels early in normal T cell development during the DN and DP stages and then at lower levels in the CD4SP and CD8SP. Heme binding to the negative regulatory protein BACH1 causes dissociation of BACH1 from the Hmox1 promoter and increased Hmox1 transcription, while expression and stability of Alas1 mRNA is under negative feedback control by heme. Therefore, we predicted that increased IFH in FLVCR-knockout thymocytes would lead to an increase in Hmox1 mRNA and a decrease in Alas1 mRNA levels. We compared expression of heme metabolic genes in FLVCR-knockout and control thymocytes. Flvcr expression was nearly absent in FLVCR-knockout DN and DP cells, however, there was a slight increase in Flvcr expression by the few CD4SP and CD8SP present. To understand this result, we analyzed the extent of genomic Flvcr deletion in FLVCR-knockout thymocytes and peripheral B and T cells by genomic qPCR. DN and DP thymocytes had near complete deletion of Flvcr while CD4SP and CD8SP had slightly less-efficient deletion, likely accounting for the increased Flvcr mRNA levels. Strikingly, Flvcr deletion in the few peripheral T cells present was 50–60% in contrast to peripheral B cells (>90%): only those T cells with incomplete Flvcr deletion survived, further underscoring the absolute requirement for FLVCR in developing T cells. We next examined Hmox1 mRNA expression and found that Hmox1 expression was higher in FLVCR-knockout DP, CD4SP, and CD8SP compared to wild-type FLVCR controls. This supports our hypothesis that FLVCR loss leads to increased IFH during T cell development. Alas1 expression was similar in FLVCR-knockout and control thymocytes, a finding that could be explained because heme regulates ALAS1 activity not only at the transcriptional level but also at the post-transcriptional level. Thus Alas1 expression may not be a good indicator of IFH. In summary, we developed a method to quantify relative free heme levels in developing thymocytes through the measurement of heme metabolic gene expression and found that IFH levels were increased in FLVCR-knockout thymocytes compared to controls. Whether and how excess free heme derails the T cell developmental program, remains to be discovered. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Comparative single-cell trajectory network enrichment identifies pseudo-temporal systems biology patterns in hematopoiesis and CD8 T-cell development

2020 ◽  
Author(s):  
Alexander G. B. Grønning ◽  
Mhaned Oubounyt ◽  
Kristiyan Kanev ◽  
Jesper Lund ◽  
Tim Kacprowski ◽  
...  
Keyword(s):  
Data Analysis ◽  
Systems Biology ◽  
T Cell ◽  
Single Cell ◽  
Expression Profiles ◽  
T Cell Development ◽  
Cd8 T Cell ◽  
Cell Development ◽  
Molecular Control ◽  
Cell Trajectory

AbstractSingle cell transcriptomics (scRNA-seq) technologies allow for investigating cellular processes on an unprecedented resolution. While software packages for scRNA-seq raw data analysis exist, no method for the extraction of systems biology signatures that drive different pseudo-time trajectories exists. Hence, pseudo-temporal molecular sub-network expression profiles remain undetermined, thus, hampering our understanding of the molecular control of cellular development on a single cell resolution. We have developed Scellnetor, the first network-constraint time-series clustering algorithm implemented as interactive webtool to identify modules of genes connected in a molecular interaction network that show differentiating temporal expression patterns. Scellnetor allows selecting two differentiation courses or two developmental trajectories for comparison on a systems biology level. Scellnetor identifies mechanisms driving hematopoiesis in mouse and mechanistically interpretable subnetworks driving dysfunctional CD8 T-cell development in chronic infections. Scellnetor is the first method to allow for single cell trajectory network enrichment for systems level hypotheses generation, thus lifting scRNA-seq data analysis to a systems biology level. It is available as an interactive online tool at https://exbio.wzw.tum.de/scellnetor/.

Dysregulation of T-Cell Development in Adrenal Glucocorticoid-Deprived Rats

2009 ◽  
Vol 234 (9) ◽  
pp. 1067-1074 ◽  
Author(s):  
Zorica Stojić-Vukanić ◽  
Aleksandra Rauški ◽  
Duško Kosec ◽  
Katarina Radojević ◽  
Ivan Pilipović ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Male Rats ◽  
Double Positive ◽  
Cell Repertoire ◽  
Single Positive ◽  
The Impact

A number of different experimental approaches have been used to elucidate the impact of basal levels of adrenal gland-derived glucocorticoids (GCs) on T cell development, and thereby T cell-mediated immune responses. However, the relevance of the adrenal GCs to T cell development is still far from clear. This study was undertaken to explore the relevance of basal levels of GCs to T cell differentiation/maturation. Eight days post-adrenalectomy in adult male rats the thymocyte yield, apoptotic and proliferative rate and the relationship amongst major thymocyte subsets, as defined by TCRαβ/CD4/CD8 expression, were examined using flow cytometry. Adrenal GC deprivation decreased thymocyte apoptosis and altered the kinetics of T cell differentiation/maturation. In the adrenalectomized rats there was increased thymic hypercellularity and an over-representation of the CD4+CD8+ double positive (DP) TCRαβlow cells entering selection, as well as increased numbers of their DP TCRαβ− immediate precursors. These changes were accompanied with under-representation of the postselected DP TCRαβhigh and the most mature CD4−CD8+ and, particularly, CD4+CD8− single positive (SP) TCRαβhigh cells. This data suggests that withdrawal of adrenal GCs produces alterations in the thymocyte selection processes, possibly affecting the diversity of functional T cell repertoire and generation of potentially self-reactive cells as indicated by the reduced proportion and number of CD4−CD8− double negative TCRαβhigh cells. In addition, it indicates that GCs influence the post-selection maturation of thymocytes and plays a regulatory role in controlling the ratio of mature CD4+CD8−/CD4−CD8+ SP TCRαβhigh cells.

Analysis of TCR, pTα, and RAG-1 in T-acute lymphoblastic leukemias improves understanding of early human T-lymphoid lineage commitment

Blood ◽  
2003 ◽  
Vol 101 (7) ◽  
pp. 2693-2703 ◽  
Author(s):  
Vahid Asnafi ◽  
Kheira Beldjord ◽  
Emmanuelle Boulanger ◽  
Béatrice Comba ◽  
Patricia Le Tutour ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Development ◽  
Cell Receptor ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Double Positive ◽  
Healthy Human ◽  
Human T Cell ◽  
Single Positive ◽  
Rag 1 ◽  
Early Human

T-acute lymphoblastic leukemias (T-ALLs) derive from human T-lymphoid precursors arrested at various early stages of development. Correlation of phenotype and T-cell receptor (TCR) status with RAG-1 and pTα transcription in 114 T-ALLs demonstrated that they largely reflect physiologic T-lymphoid development. Half the TCRαβ lineage T-ALLs expressed a pre-TCR, as evidenced by RAG-1, pTα, and cTCRβ expression, absence of TCRδ deletion, and a sCD3−, CD1a+, CD4/8 double-positive (DP) phenotype, in keeping with a population undergoing β selection. Most TCRγδ T-ALLs were pTα, terminal deoxynucleotidyl transferase (TdT), and RAG-1lo/neg, double-negative/single-positive (DN/SP), and demonstrated only TCRβ DJ rearrangement, whereas 40% were pTα, TdT, and RAG-1 positive, DP, and demonstrated TCRβ V(D)J rearrangement, with cTCRβ expression in proportion. As such they may correspond to TCRαβ lineage precursors selected by TCRγδ expression, to early γδ cells recently derived from a pTα+ common αβ/γδ precursor, or to a lineage-deregulated αβ/γδ intermediate. Approximately 30% of T-ALLs were sCD3/cTCRβ− and corresponded to nonrestricted thymic precursors because they expressed non–T-restricted markers such as CD34, CD13, CD33, and CD56 and were predominantly DN, CD1a, pTα, and RAG-1 low/negative, despite immature TCRδ and TCRγ rearrangements. TCR gene configuration identified progressive T-lymphoid restriction. T-ALLs, therefore, provide homogeneous expansions of minor human lymphoid precursor populations that can aid in the understanding of healthy human T-cell development.

scholarly journals The role of the Runx transcription factors in thymocyte differentiation and in homeostasis of naive T cells

2007 ◽  
Vol 204 (8) ◽  
pp. 1945-1957 ◽  
Author(s):  
Takeshi Egawa ◽  
Robert E. Tillman ◽  
Yoshinori Naoe ◽  
Ichiro Taniuchi ◽  
Dan R. Littman
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Cytotoxic T Cell ◽  
Thymocyte Development ◽  
Double Positive ◽  
Interleukin 7 ◽  
Genes Encoding ◽  
Single Positive

Members of the Runx family of transcriptional regulators are required for the appropriate expression of CD4 and CD8 at discrete stages of T cell development. The roles of these factors in other aspects of T cell development are unknown. We used a strategy to conditionally inactivate the genes encoding Runx1 or Runx3 at different stages of thymocyte development, demonstrating that Runx1 regulates the transitions of developing thymocytes from the CD4−CD8− double-negative stage to the CD4+CD8+ double-positive (DP) stage and from the DP stage to the mature single-positive stage. Runx1 and Runx3 deficiencies caused marked reductions in mature thymocytes and T cells of the CD4+ helper and CD8+ cytotoxic T cell lineages, respectively. Runx1-deficient CD4+ T cells had markedly reduced expression of the interleukin 7 receptor and exhibited shorter survival. In addition, inactivation of both Runx1 and Runx3 at the DP stages resulted in a severe block in development of CD8+ mature thymocytes. These results indicate that Runx proteins have important roles at multiple stages of T cell development and in the homeostasis of mature T cells.

scholarly journals The Metabolic Landscape of Thymic T Cell Development In Vivo and In Vitro

2021 ◽  
Vol 12 ◽  
Author(s):  
Victoria Sun ◽  
Mark Sharpley ◽  
Karolina E. Kaczor-Urbanowicz ◽  
Patrick Chang ◽  
Amélie Montel-Hagen ◽  
...  
Keyword(s):  
T Cell ◽  
Oxidative Phosphorylation ◽  
Metabolic Pathways ◽  
T Cell Development ◽  
Cell Development ◽  
Mouse Bone Marrow ◽  
Double Positive ◽  
Human T Cell

Although metabolic pathways have been shown to control differentiation and activation in peripheral T cells, metabolic studies on thymic T cell development are still lacking, especially in human tissue. In this study, we use transcriptomics and extracellular flux analyses to investigate the metabolic profiles of primary thymic and in vitro-derived mouse and human thymocytes. Core metabolic pathways, specifically glycolysis and oxidative phosphorylation, undergo dramatic changes between the double-negative (DN), double-positive (DP), and mature single-positive (SP) stages in murine and human thymus. Remarkably, despite the absence of the complex multicellular thymic microenvironment, in vitro murine and human T cell development recapitulated the coordinated decrease in glycolytic and oxidative phosphorylation activity between the DN and DP stages seen in primary thymus. Moreover, by inducing in vitro T cell differentiation from Rag1-/- mouse bone marrow, we show that reduced metabolic activity at the DP stage is independent of TCR rearrangement. Thus, our findings suggest that highly conserved metabolic transitions are critical for thymic T cell development.

scholarly journals Defective development of thymocytes overexpressing the costimulatory molecule, heat-stable antigen.

1994 ◽  
Vol 179 (1) ◽  
pp. 177-184 ◽  
Author(s):  
M R Hough ◽  
F Takei ◽  
R K Humphries ◽  
R Kay
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Costimulatory Molecule ◽  
Cell Development ◽  
Double Negative ◽  
Double Positive ◽  
Heat Stable ◽  
Heat Stable Antigen ◽  
Single Positive

Heat-stable antigen (HSA) is a small, glycosyl phosphatidylinositol-anchored protein that can act as a costimulatory molecule for antigen-dependent activation of helper T cells. In addition to being expressed on antigen-presenting B cells, HSA is also expressed during the initial stages of T cell development in the thymus. HSA levels are very high on immature CD4-, CD8- double negative thymocytes, but are reduced on CD4+, CD8+ double positive cells undergoing selection in the thymus, and are entirely eliminated when these cells differentiate into immunologically competent CD4+ or CD8+ single positive T cells. To examine the potential roles of this molecule in T cell development and selection, we generated transgenic mice in which HSA was highly expressed on all classes of thymocytes. The consequence of deregulated HSA expression was a pronounced reduction in the numbers of double positive and single positive thymocytes, whereas the numbers of their double negative precursors were largely unaffected. These results demonstrate that downregulation of HSA expression at the double positive stage is a critical event in thymocyte development. The depletion of thymocytes resulting from HSA overexpression begins at the same time as the onset of negative selection, suggesting that HSA may provide signals that contribute to determining the efficiency of this process.

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