An Intrinsic Mechanism Predisposes Foxp3-Expressing Regulatory T Cells to Th2 Conversion In Vivo

Naturally occurring regulatory T (nTreg) cells express Foxp3 and were originally discovered as immune suppressors critical for self-tolerance and immune homeostasis. Through yet-to-be-defined mechanisms, nTreg cells were recently shown to convert into proinflammatory cells. Particularly, attenuation of Foxp3 expression led to Th2 conversion of nTreg cells in vivo. In this paper, we demonstrated an nTreg-specific mechanism controlling their Th2 conversion. We found that wild-type nTreg cells expressing reduced levels of Foxp3 but not those expressing no Foxp3 produced the Th2 cytokine IL-4. Intriguingly, IL-4 production by converted nTreg cells is required for Th2 differentiation of coexisting naive CD4 T cells in vivo, suggesting that Th2 conversion of nTreg cells might be critical for directing Th2 immune responses. Th2 conversion of nTreg cells was not due to their inability to become Th1 cells, because IFN-γ was produced by Foxp3-low–expressing cells when IL-4/STAT-6 signaling was abrogated. Surprisingly, however, unlike naive CD4 T cells whose IL-4 production is dependent on STAT-6, Foxp3-low–expressing cells generated IL-4 independent of STAT-6, indicating an intrinsic mechanism that favors nTreg-to-Th2 differentiation. Indeed, compared with naive CD4 T cells, nTreg expressed elevated levels of GATA-3 independent of STAT-6. And GATA-3 was required for nTreg-to-Th2 conversion. Foxp3 may account for this GATA-3 upregulation in nTreg cells, because ectopic expression of Foxp3 preferentially promoted GATA-3 but not T-bet expression. Thus, we have identified an intrinsic mechanism that imposes a Th2/Th1 imbalance and predisposes Foxp3-expressing cells to IL-4 production independent of STAT-6 signaling

Regulation of GATA-3 Expression during CD4 Lineage Differentiation

GATA-3 is necessary for the development of MHC class II-restricted CD4 T cells, and its expression is increased during positive selection of these cells. TCR signals drive this upregulation, but the signaling pathways that control this process are not well understood. Using genetic and pharmacological approaches, we show that GATA-3 upregulation during thymocyte-positive selection is the result of additive inputs from the Ras/MAPK and calcineurin pathways. This upregulation requires the presence of the transcription factor c-Myb. Furthermore, we show that TH-POK can also upregulate GATA-3 in double-positive thymocytes, suggesting the existence of a positive feedback loop that contributes to lock in the initial commitment to the CD4 lineage during differentiation

SATB1 Defines the Developmental Context for Gene Silencing by Xist in Lymphoma and Embryonic Cells

SummaryThe noncoding Xist RNA triggers silencing of one of the two female X chromosomes during X inactivation in mammals. Gene silencing by Xist is restricted to a special developmental context in early embryos and specific hematopoietic precursors. Here, we show that Xist can initiate silencing in a lymphoma model. We identify the special AT-rich binding protein SATB1 as an essential silencing factor. Loss of SATB1 in tumor cells abrogates the silencing function of Xist. In lymphocytes Xist localizes along SATB1-organized chromatin and SATB1 and Xist influence each other's pattern of localization. SATB1 and its homolog SATB2 are expressed during the initiation window for X inactivation in ES cells. Importantly, viral expression of SATB1 or SATB2 enables gene silencing by Xist in embryonic fibroblasts, which normally do not provide an initiation context. Thus, our data establish SATB1 as a crucial silencing factor contributing to the initiation of X inactivation

Transcription factor YY1 is essential for regulation of the Th2 cytokine locus and for Th2 cell differentiation

The Th2 locus control region (LCR) has been shown to be important in efficient and coordinated cytokine gene regulation during Th2 cell differentiation. However, the molecular mechanism for this is poorly understood. To study the molecular mechanism of the Th2 LCR, we searched for proteins binding to it. We discovered that transcription factor YY1 bound to the LCR and the entire Th2 cytokine locus in a Th2-specific manner. Retroviral overexpression of YY1 induced Th2 cytokine expression. CD4-specific knockdown of YY1 in mice caused marked reduction in Th2 cytokine expression, repressed chromatin remodeling, decreased intrachromosomal interactions, and resistance in an animal model of asthma. YY1 physically associated with GATA-binding protein-3 (GATA3) and is required for GATA3 binding to the locus. YY1 bound to the regulatory elements in the locus before GATA3 binding. Thus, YY1 cooperates with GATA3 and is required for regulation of the Th2 cytokine locus and Th2 cell differentiation

Gata3 Acts Downstream of β-Catenin Signaling to Prevent Ectopic Metanephric Kidney Induction

Metanephric kidney induction critically depends on mesenchymal–epithelial interactions in the caudal region of the nephric (or Wolffian) duct. Central to this process, GDNF secreted from the metanephric mesenchyme induces ureter budding by activating the Ret receptor expressed in the nephric duct epithelium. A failure to regulate this pathway is believed to be responsible for a large proportion of the developmental anomalies affecting the urogenital system. Here, we show that the nephric duct-specific inactivation of the transcription factor gene Gata3 leads to massive ectopic ureter budding. This results in a spectrum of urogenital malformations including kidney adysplasia, duplex systems, and hydroureter, as well as vas deferens hyperplasia and uterine agenesis. The variability of developmental defects is reminiscent of the congenital anomalies of the kidney and urinary tract (CAKUT) observed in human. We show that Gata3 inactivation causes premature nephric duct cell differentiation and loss of Ret receptor gene expression. These changes ultimately affect nephric duct epithelium homeostasis, leading to ectopic budding of interspersed cells still expressing the Ret receptor. Importantly, the formation of these ectopic buds requires both GDNF/Ret and Fgf signaling activities. We further identify Gata3 as a central mediator of β-catenin function in the nephric duct and demonstrate that the β-catenin/Gata3 pathway prevents premature cell differentiation independently of its role in regulating Ret expression. Together, these results establish a genetic cascade in which Gata3 acts downstream of β-catenin, but upstream of Ret, to prevent ectopic ureter budding and premature cell differentiation in the nephric duct

Oncogenic role of Pax5 in the T-lymphoid lineage upon ectopic expression from the immunoglobulin heavy-chain locus

Four of 9 PAX transcription factor genes have been associated with chromosomal translocations in human tumors, although their oncogenic potential has not yet been demonstrated in transgenic mouse models. The B-lymphoidPAX5 gene participates in the generation of the t(9;14)(p13;q32) translocation in germinal center B cells, which leads to deregulated PAX5 expression under the control of the immunoglobulin heavy-chain (IgH) locus in a subset of B-cell non-Hodgkin lymphomas. Here we reconstructed a human t(9;14) translocation in a knock-in mouse by inserting a PAX5 minigene into the IgH locus. The IgH(P5ki) allele, which corresponds to a germline rather than somatic mutation, is activated in multipotent hematopoietic progenitors and is subsequently expressed in dendritic cells (DCs) and in natural killer (NK), T, and B cells. Ectopic Pax5 expression interferes with normal T-cell development and causes immature T-lymphoblastic lymphomas in IgH(P5ki/+) and IgH(P5ki/P5ki) mice. Aggressive T-cell lymphomas develop even faster in Ik(Pax5/+) mice expressing Pax5 from the Ikaros locus. Pax5 expression in thymocytes activates B-cell-specific genes and represses T-lymphoid genes, suggesting that Pax5 contributes to lymphomagenesis by deregulating the T-cell gene-expression program. These data identify Pax5 as a potent oncogene and demonstrate that the T-lymphoid lineage is particularly sensitive to the oncogenic action of Pax5

Oncogenic role of Pax5 in the T-lymphoid lineage upon ectopic expression from the immunoglobulin heavy-chain locus

Four of 9 PAX transcription factor genes have been associated with chromosomal translocations in human tumors, although their oncogenic potential has not yet been demonstrated in transgenic mouse models. The B-lymphoidPAX5 gene participates in the generation of the t(9;14)(p13;q32) translocation in germinal center B cells, which leads to deregulated PAX5 expression under the control of the immunoglobulin heavy-chain (IgH) locus in a subset of B-cell non-Hodgkin lymphomas. Here we reconstructed a human t(9;14) translocation in a knock-in mouse by inserting a PAX5 minigene into the IgH locus. The IgH(P5ki) allele, which corresponds to a germline rather than somatic mutation, is activated in multipotent hematopoietic progenitors and is subsequently expressed in dendritic cells (DCs) and in natural killer (NK), T, and B cells. Ectopic Pax5 expression interferes with normal T-cell development and causes immature T-lymphoblastic lymphomas in IgH(P5ki/+) and IgH(P5ki/P5ki) mice. Aggressive T-cell lymphomas develop even faster in Ik(Pax5/+) mice expressing Pax5 from the Ikaros locus. Pax5 expression in thymocytes activates B-cell-specific genes and represses T-lymphoid genes, suggesting that Pax5 contributes to lymphomagenesis by deregulating the T-cell gene-expression program. These data identify Pax5 as a potent oncogene and demonstrate that the T-lymphoid lineage is particularly sensitive to the oncogenic action of Pax5

Nephric lineage specification by Pax2 and Pax8

The mammalian kidney develops in three successive steps from the initial pronephros via the mesonephros to the adult metanephros. Although the nephric lineage is specified during pronephros induction, no single regulator, including the transcription factor Pax2 or Pax8, has yet been identified to control this initial phase of kidney development. In this paper, we demonstrate that mouse embryos lacking both Pax2 and Pax8 are unable to form the pronephros or any later nephric structures. In these double-mutant embryos, the intermediate mesoderm does not undergo the mesenchymal-epithelial transitions required for nephric duct formation, fails to initiate the kidney-specific expression of Lim1 and c-Ret, and is lost by apoptosis 1 d after failed pronephric induction. Conversely, retroviral misexpression of Pax2 was sufficient to induce ectopic nephric structures in the intermediate mesoderm and genital ridge of chick embryos. Together, these data identify Pax2 and Pax8 as critical regulators that specify the nephric lineage