Expression data from sorted Id3-GFP hi Id2-YFP int and Id3-GFP lo Id2-YFP hi activated CD8 T cells
ABSTRACT: During an immune response, CD8 T cells fall along a gradient of memory potential, but the regulators of these fate decsisions are not well understood. We utlized Id3-GFP and Id2-YFP reporter mice to elucidate the role of Id3 and Id2 during early CD8 T cell differentiation by gene expression. Id3-GFP hi Id2-YFP int or Id3-GFP lo Id2-YFP hi OT-I cells were sorted into trizol at day 6 of VSV-OVA infection and analyzed by microarray
Project description:During an immune response, CD8 T cells fall along a gradient of memory potential, but the regulators of these fate decsisions are not well understood. We utlized Id3-GFP and Id2-YFP reporter mice to elucidate the role of Id3 and Id2 during early CD8 T cell differentiation by gene expression. Overall design: Id3-GFP hi Id2-YFP int or Id3-GFP lo Id2-YFP hi OT-I cells were sorted into trizol at day 6 of VSV-OVA infection and analyzed by microarray
Project description:During infection, naive CD8(+) T cells differentiate into effector cells, which are armed to eliminate pathogens, and memory cells, which are poised to protect against reinfection. The transcriptional program that regulates terminal differentiation into short-lived effector-memory versus long-lived memory cells is not clearly defined. Through the use of mice expressing reporters for the DNA-binding inhibitors Id2 and Id3, we identified Id3(hi) precursors of long-lived memory cells before the peak of T cell population expansion or upregulation of cell-surface receptors that indicate memory potential. Deficiency in Id2 or Id3 resulted in loss of distinct CD8(+) effector and memory populations, which demonstrated unique roles for these inhibitors of E-protein transcription factors. Furthermore, cytokines altered the expression of Id2 and Id3 differently, which provides insight into how external cues influence gene expression.
Project description:Regulatory T (Treg) cells suppress the development of inflammatory disease, but our knowledge of transcriptional regulators that control this function remains incomplete. Here we show that expression of Id2 and Id3 in Treg cells was required to suppress development of fatal inflammatory disease. We found that T cell antigen receptor (TCR)-driven signaling initially decreased the abundance of Id3, which led to the activation of a follicular regulatory T (TFR) cell-specific transcription signature. However, sustained lower abundance of Id2 and Id3 interfered with proper development of TFR cells. Depletion of Id2 and Id3 expression in Treg cells resulted in compromised maintenance and localization of the Treg cell population. Thus, Id2 and Id3 enforce TFR cell checkpoints and control the maintenance and homing of Treg cells.
Project description:Inhibitor of DNA binding (Id) proteins, including Id1-4, are transcriptional regulators involved in promoting cell proliferation and survival in various cell types. Although upregulation of Id proteins is associated with a broad spectrum of tumors, recent studies have identified that Id3 plays a tumor-suppressor role in the development of Burkitt's lymphoma in humans and hepatosplenic T cell lymphomas in mice. In this article, we report rapid lymphoma development in Id2/Id3 double-knockout mice that is caused by unchecked expansion of invariant NKT (iNKT) cells or a unique subset of innate-like CD1d-independent T cells. These populations began to expand in neonatal mice and, upon malignant transformation, resulted in mortality between 3 and 11 mo of age. The malignant cells also gave rise to lymphomas upon transfer to Rag-deficient and wild-type hosts, reaffirming their inherent tumorigenic potential. Microarray analysis revealed a significantly modified program in these neonatal iNKT cells that ultimately led to their malignant transformation. The lymphoma cells demonstrated chromosome instability along with upregulation of several signaling pathways, including the cytokine-cytokine receptor interaction pathway, which can promote their expansion and migration. Dysregulation of genes with reported driver mutations and the NF-?B pathway were found to be shared between Id2/Id3 double-knockout lymphomas and human NKT tumors. Our work identifies a distinct premalignant state and multiple tumorigenic pathways caused by loss of function of Id2 and Id3. Thus, conditional deletion of Id2 and Id3 in developing T cells establishes a unique animal model for iNKT and relevant innate-like lymphomas.
Project description:The innate-like T cells expressing V?1.1 and V?6.3 represent a unique T cell lineage sharing features with both the ?? T and the invariant NKT cells. The population size of V?1.1(+)V?6.3(+) T cells is tightly controlled and usually contributes to a very small proportion of thymic output, but the underlying mechanism remains enigmatic. Deletion of Id3, an inhibitor of E protein transcription factors, can induce an expansion of the V?1.1(+)V?6.3(+) T cell population. This phenotype is much stronger on the C57BL/6 background than on the 129/sv background. Using quantitative trait linkage analysis, we identified Id2, a homolog of Id3, to be the major modifier of Id3 in limiting V?1.1(+)V?6.3(+) T cell expansion. The V?1.1(+)V?6.3(+) phenotype is attributed to an intrinsic weakness of Id2 transcription from Id2 C57BL/6 allele, leading to an overall reduced dosage of Id proteins. However, complete removal of both Id2 and Id3 genes in developing T cells suppressed the expansion of V?1.1(+)V?6.3(+) T cells because of decreased proliferation and increased cell death. We showed that conditional knockout of Id2 alone is sufficient to promote a moderate expansion of ?? T cells. These regulatory effects of Id2 and Id3 on V?1.1(+)V?6.3(+) T cells are mediated by titration of E protein activity, because removing one or more copies of E protein genes can restore V?1.1(+)V?6.3(+) T cell expansion in Id2 and Id3 double conditional knockout mice. Our data indicated that Id2 and Id3 collaboratively control survival and expansion of the ?? lineage through modulating a proper threshold of E proteins.
Project description:Gene knockout experiments in mice have suggested a hierarchical model of early B cell commitment wherein E2A proteins (E47 and E12) activate early B cell factor (Ebf1), which in turn activates expression of the B cell commitment factor, Pax5. In IL-7 receptor alpha (IL-7Ralpha) knockout mice, B cell development is blocked before B-lineage commitment at the prepro-B cell stage in adult animals. In IL-7Ralpha(-/-) prepro-B cells, E47 is expressed and yet is insufficient to transcriptionally activate the putative downstream target gene, Ebf1. In this study, we show that further increases of E47 expression in IL-7Ralpha(-/-) prepro-B cells fails to activate Ebf1, but rather leads to a dramatic induction of the E2A inhibitory factors, Id2 and Id3. In contrast, enforced expression of Ebf1 in IL-7Ralpha(-/-) bone marrow potently down-regulates Id2 and Id3 mRNA expression and restores B cell differentiation in vivo. Down-regulation of both Id2 and Id3 during B cell specification is essential in that overexpression of either Id2 or Id3 in wild-type bone marrow blocks B cell specification at the prepro-B cell stage. Collectively, these studies suggest a model where Ebf1 induction specifies the B cell fate by dramatically increasing activity of E47 at the posttranslational level.
Project description:The invariant NKT (iNKT) cells represent a unique group of ?? T cells that have been classified based on their exclusive usage of the invariant V?14J?18 TCR?-chain and their innate-like effector function. Thus far, the transcriptional programs that control V?14J?18 TCR? rearrangements and the population size of iNKT cells are still incompletely defined. E protein transcription factors have been shown to play necessary roles in the development of multiple T cell lineages, including iNKT cells. In this study, we examined E protein functions in T cell development through combined deletion of genes encoding E protein inhibitors Id2 and Id3. Deletion of Id2 and Id3 in T cell progenitors resulted in a partial block at the pre-TCR selection checkpoint and a dramatic increase in numbers of iNKT cells. The increase in iNKT cells is accompanied with a biased rearrangement involving V?14 to J?18 recombination at the double-positive stage and enhanced proliferation of iNKT cells. We further demonstrate that a 50% reduction of E proteins can cause a dramatic switch from iNKT to innate-like ?? T cell fate in Id2- and Id3-deficient mice. Collectively, these findings suggest that Id2- and Id3-mediated inhibition of E proteins controls iNKT development by restricting lineage choice and population expansion.
Project description:Transforming growth factors beta (TGF-betas) inhibit growth of epithelial cells and induce differentiation changes, such as epithelial-mesenchymal transition (EMT). On the other hand, bone morphogenetic proteins (BMPs) weakly affect epithelial cell growth and do not induce EMT. Smad4 transmits signals from both TGF-beta and BMP pathways. Stimulation of Smad4-deficient epithelial cells with TGF-beta 1 or BMP-7 in the absence or presence of exogenous Smad4, followed by cDNA microarray analysis, revealed 173 mostly Smad4-dependent, TGF-beta-, or BMP-responsive genes. Among 25 genes coregulated by both factors, inhibitors of differentiation Id2 and Id3 showed long-term repression by TGF-beta and sustained induction by BMP. The opposing regulation of Id genes is critical for proliferative and differentiation responses. Hence, ectopic Id2 or Id3 expression renders epithelial cells refractory to growth inhibition and EMT induced by TGF-beta, phenocopying the BMP response. Knockdown of endogenous Id2 or Id3 sensitizes epithelial cells to BMP, leading to robust growth inhibition and induction of transdifferentiation. Thus, Id genes sense Smad signals and create a permissive or refractory nuclear environment that defines decisions of cell fate and proliferation.