E2A and HEB Are Required to Block Thymocyte Proliferation Prior to Pre-TCR Expression

Wojciechowski, Jason; Lai, Anne Y.; Kondo, Motonari; Zhuang, Yuan

doi:10.4049/jimmunol.178.9.5717

Cited by 101 publications

(130 citation statements)

References 43 publications

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“…HEB −/− T cells possess compromised Notch1 function and lose T cell potential [37]. The activity of E proteins is crucial for the DN2-to- DN3 transition and TCR gene rearrangement; loss of E2A leads to a reverse differentiation from DN3 cells to DN2 cells [38]. E proteins lose their function under high levels of Id1 protein, which can inhibit the DNA-binding activity of E proteins.…”

Section: Discussionmentioning

confidence: 99%

Downregulation of the transcription factor KLF4 is required for the lineage commitment of T cells

et al. 2011

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The roles of the reprogramming factors Oct4, Sox2, c-Myc and Klf4 in early T cell development are incompletely defined. Here, we show that Klf4 is the only reprogramming factor whose expression is downregulated when early thymic progenitors (ETPs) differentiate into T cells. Enforced expression of Klf4 in uncommitted progenitors severely impaired T cell development mainly at the DN2-to-DN3 transition when T cell lineage commitment occurs and affected the transcription of a variety of genes with crucial functions in early T cell development, including genes involved in microenvironmental signaling (IL-7Rα), Notch target genes (Deltex1), and essential T cell lineage regulatory or inhibitory genes (Bcl11a, SpiB, and Id1). The survival of thymocytes and the rearrangement at the Tcrb locus were impaired in the presence of enforced Klf4 expression. The defects in the DN1-to-DN2 and DN2-to-DN3 transitions in Klf4 transgenic mice could not be rescued by the introduction of a TCR transgene, but was partially rescued by restoring the expression of IL-7Rα. Thus, our data indicate that the downregulation of Klf4 is a prerequisite for T cell lineage commitment.

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Section: Discussionmentioning

confidence: 99%

Downregulation of the transcription factor KLF4 is required for the lineage commitment of T cells

et al. 2011

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“…Data are relative to the 0-h sample, whose value was set at 100 and is representative of two independent experiments. stage (Wojciechowski et al 2007). The genotypes of the floxed mice were either Tcf12…”

Section: Dn3 Cells From Rag2mentioning

confidence: 99%

“…LckCre + mice were generated as described (Pan et al 1999;Zhang et al 1999;Wojciechowski et al 2007). All animal experiments were performed according to guidelines from the National Institutes of Health and with an approved protocol from the University of Pennsylvania Animal Care and Use Committee.…”

Section: Micementioning

confidence: 99%

Pre-TCR signaling inactivates Notch1 transcription by antagonizing E2A

Yashiro–Ohtani¹,

He²,

Ohtani³

et al. 2009

Genes Dev.

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Precise control of the timing and magnitude of Notch signaling is essential for the normal development of many tissues, but the feedback loops that regulate Notch are poorly understood. Developing T cells provide an excellent context to address this issue. Notch1 signals initiate T-cell development and increase in intensity during maturation of early T-cell progenitors (ETP) to the DN3 stage. As DN3 cells undergo β-selection, during which cells expressing functionally rearranged TCRβ proliferate and differentiate into CD4+CD8+ progeny, Notch1 signaling is abruptly down-regulated. In this report, we investigate the mechanisms that control Notch1 expression during thymopoiesis. We show that Notch1 and E2A directly regulate Notch1 transcription in pre-β-selected thymocytes. Following successful β-selection, pre-TCR signaling rapidly inhibits Notch1 transcription via signals that up-regulate Id3, an E2A inhibitor. Consistent with a regulatory role for Id3 in Notch1 down-regulation, post-β-selected Id3-deficient thymocytes maintain Notch1 transcription, whereas enforced Id3 expression decreases Notch1 expression and abrogates Notch1-dependent T-cell survival. These data provide new insights into Notch1 regulation in T-cell progenitors and reveal a direct link between pre-TCR signaling and Notch1 expression during thymocyte development. Our findings also suggest new strategies for inhibiting Notch1 signaling in pathologic conditions.

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“…In contrast to B cell development, where E2A plays a predominant role, T cell development is regulated by the combined dosage of E2A and HEB (21). Thus, conditional knock-out of both E2A and Heb genes is required to induce severe developmental defects in the T cell lineage (22,23). It has been shown that LckCre-mediated conditional knock-out of E2A and Heb at the double-negative (DN; CD4 Ϫ CD8 Ϫ ) stage of thymocyte development blocks further development of the DN cells into the double-positive (DP; CD4 ϩ CD8 ϩ ) stage (22).…”

Section: Resultsmentioning

confidence: 99%

Generation and Analysis of Partially Haploid Cells with Cre-mediated Chromosome Deletion in the Lymphoid System

Zhu

Kim

et al. 2010

Journal of Biological Chemistry

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The fast accumulation of mutant mouse strains in recent years has provided an invaluable resource for phenotype-based genetic screens. However, study of lymphoid phenotypes can be obscured or impractical if homozygous mutations cause early embryonic defects. To aid phenotype screening of germ line mutations in the lymphoid system, we developed a method to induce loss of heterozygosity (LOH) in developing lymphocytes through chromosome deletion. Chromosome deletion was triggered by Cre/loxP-mediated inverse sister chromatid recombination in the G 2 /M phase of the cell cycle, leading to the generation of daughter cells missing part of or the entire recombinant chromosome. We show that the resulting cells were viable and capable of additional rounds of cell division, thus providing raw materials for subsequent phenotypic assessment. We used the recombination system to induce LOH at the E2A locus in developing B cells. A significant loss of pro-B and pre-B cells was observed when the wild-type allele was removed by chromosome deletion from the E2A heterozygous mice, a result consistent with the required role for E2A in B cell development. We also demonstrated the effectiveness of Cre-mediated chromosome deletion in the LOH assay for HEB function in T cell development. Thus, the Cre-mediated chromosome deletion provides a new and effective method for genome-wide assessment of germ line mutations in the lymphoid system. Over 4000 protein-coding genes have been mutated and studied in the mouse by gene targeting, gene trapping, or other mutagenic methods in the past 2 decades (1, 2). This number is expected to increase dramatically following the recent calls for genome-scale coverage of germ line mutations by several international consortia (3). Studies of the existing germ line mutations have implied that approximately one-third of null mutations cause embryonic or neonatal lethality (4). Consequently, the effect of germ line mutations on postnatal life cannot be easily assessed for the genes playing important roles in both embryonic and postnatal life. Further analysis of tissue-specific gene function often requires a completely independent effort to establish a new allele for Cre/ lox-mediated conditional gene knock-out. Although the Cre/lox system can effectively address the embryonic problem and permit the study of tissue-specific gene function, the experimental system is not practical for large-scale genetic analysis. Thus far, there is no simple method available for systematic evaluation of lymphoid phenotypes of lethal mutations accumulated from a large-scale mutagenesis approach.Mitotic recombination provides one possible way to assess somatic phenotypes of pre-existing mutations. A recombination between homologous chromosomes during mitosis can lead to clonal segregation of heterologous alleles. The mosaics resulting from mitotic recombination allow for functional analysis of homozygous clones in an otherwise heterozygous background (5, 6). The feasibility of mosaic analysis in mice was demonstrated in the lo...

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E2A and HEB Are Required to Block Thymocyte Proliferation Prior to Pre-TCR Expression

Cited by 101 publications

References 43 publications

Downregulation of the transcription factor KLF4 is required for the lineage commitment of T cells

Downregulation of the transcription factor KLF4 is required for the lineage commitment of T cells

Pre-TCR signaling inactivates Notch1 transcription by antagonizing E2A

Generation and Analysis of Partially Haploid Cells with Cre-mediated Chromosome Deletion in the Lymphoid System

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