Conditional deletion of Gata3 shows its essential function in TH1-TH2 responses

Zhu, Jinfang; Min, Booki; Hu‐Li, Jane; Watson, Cynthia; Grinberg, Alex; Wang, Qi; Killeen, Nigel; Urban, Joseph F.; Guo, Liying; Paul, William E.

doi:10.1038/ni1128

Cited by 564 publications

(599 citation statements)

References 48 publications

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“…However, IL-4 was blocked, as demonstrated by inhibition of both IL-4 mRNA and serum IgE elevations, when B7 costimulation was abrogated. These studies thus demonstrate differential regulation of IL-4 and IL-13 in T cells by costimulatory molecules during the in vivo immune response, consistent with previous studies indicating that these Th2 cytokines can be differentially regulated by certain transcription factors [10,11]. Our studies further indicate that DX5 + cells are also a major source of IL-13.…”

Section: Discussionsupporting

confidence: 91%

“…However, other studies suggest that under certain circumstances IL-4 and IL-13 are regulated independently, and certain signaling pathways involving c-maf [10] and GATA-3 [11] can differentially regulate these two cytokines. Consistent with these findings, several in vivo studies have recently identified IL-4-independent elevations of IL-13 associated with an IFN-c-dominant response [8,12], although the cell surface or secreted molecules that induce the development of IL-13-expressing cells in this context remain uncertain.…”

Section: Introductionmentioning

confidence: 99%

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IL‐18 stimulates IL‐13‐mediated IFN‐γ‐sensitive host resistance in vivo

Liu

Whitmire

et al. 2006

Eur J Immunol

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IL-4 and IL-13 are up-regulated during in vivo responses to many nematode parasites, but increasing evidence suggests that increases in IL-13 can also occur independently of the IL-4-dominant Th2 response. Blocking B7 after Trichuris muris inoculation inhibits resistance and IL-4 elevations, instead resulting in an IFN-c-dominant response associated with susceptibility. However, blocking IFN-c under these conditions restores IL-13-dependent resistance. In this study, we examined the mechanism of IL-13 upregulation and associated protection during this in vivo immune response. CD4 + T cells and DX5 + TCR -cells were identified as the major producers of IL-13, and the DX5 + TCRcells were phenotyped as NK cells, since they expressed CD11b, IL-2Rb and Ly49C but not c-kit or FceRI. NK cell-derived IL-13 elevations were T cell-dependent, as CD4 + T cell depletion blocked IL-13 production by mesenteric lymph node cells and induced susceptibility. IL-13 expression was increased independently of IL-12; however, blocking IL-18 function inhibited IL-13 production and increased susceptibility. These results indicate that CD4 + T cells and NK cells are the major sources of IL-13 during the in vivo Th1 response induced by B7 blockade and that under these conditions, IL-18 is specifically required for the in vivo up-regulation of IL-13 production and associated host protection. IntroductionThe host protective response that develops following infection varies greatly with the particular pathogen. The type 2 immune response, associated with high levels of IL-4, IL-13 and other Th2 cytokines, provides protection against intestinal nematode parasites [1-3], while type 1 immunity, associated with an IFNc-dominant response, mediates resistance against many viruses and bacteria [4,5]. The events that lead to the development of type 1 and type 2 immunity are generally thought to be distinct, and as each response matures, production of characteristic cytokines can down-regulate the reciprocal response, resulting in further polarization.Typically, type 2 responses leading to resistance require the development of IL-4-producing effector CD4 + T cells, which then utilize autocrine IL-4 for their rapid expansion [3]. These Th2 cells mediate worm expulsion through their production of Th2 cytokines, with IL-4 and IL-13 being particularly important for the expulsion of gastrointestinal nematode parasites [1,2]. The development and expansion of IL-4-producing T cells is preferentially dependent on B7 costimulatory molecules [6,7], and in a number of infectious diseases, B7 blockade can actually deviate a Th2 response to an IFN-c-dominant Th1 response [8,9]. In the Th2 cell differentiation model just described, blockade of IL-4 production with B7 antagonists would be expected to also inhibit IL-13 production. However, other studies suggest that under certain circumstances IL-4 and IL-13 are regulated independently, and certain signaling pathways involving c-maf [10] and can differentially regulate these two cytokines. Consistent with these fin...

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Section: Discussionsupporting

confidence: 91%

Section: Introductionmentioning

confidence: 99%

IL‐18 stimulates IL‐13‐mediated IFN‐γ‐sensitive host resistance in vivo

Liu

Whitmire

et al. 2006

Eur J Immunol

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“…Silencing of CD4 resulted in the expected impairment of antigen-dependent T-cell activation [13][14][15]. siRNA-mediated knockdown of the Th2 transcription factor GATA-3 abolished the expression of its target genes, the prototypic Th2 cytokines IL-4 and IL-5, as reported for GATA-3 knockout mice [17,18]. Noteworthy, the use of stabilized siRNA proved to be absolutely mandatory since conventional siRNA identical in sequence and tested for efficiency in cell lines turned out to be largely ineffective in these assays.…”

Section: Discussionmentioning

confidence: 93%

“…We chose GATA-3 as a target given that this transcription factor is critical for the development of Th2 cells from naïve T cells, which occurs in the first days after stimulation [16][17][18]. T cells were transfected with stabilized or conventional GATA-3 siRNA, a control siRNA (siGFP) or were nucleofected without siRNA.…”

Section: Sirna-transfected T Cells Remain Functional and Maintain Knomentioning

confidence: 99%

siRNA stabilization prolongs gene knockdown in primary T lymphocytes

et al. 2008

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RNA interference (RNAi)-mediated knockdown of target gene expression represents a powerful approach for functional genomics and therapeutic applications. However, for T lymphocytes, central regulators of immunity and immunopathologies, the application of RNAi has been limited due to the lack of efficient small interfering RNA (siRNA) delivery protocols, and an inherent inefficiency of the RNAi machinery itself. Here, we use nucleofection, an optimized electroporation approach, to deliver siRNA into primary T lymphocytes with high efficiency and negligible impairment of cell function. We identify siRNA stability within the cells as the critical parameter for efficient RNAi in primary T cells. While generally short-lived and immediately lost upon T-cell activation when conventional siRNA is used, target gene knockdown becomes insensitive to cell activation and can persist for up to 2 wk in non-dividing cells with siRNA stabilized by chemical modifications. Targeting CD4 and the transcription factor GATA-3, we show that the use of stabilized siRNA is imperative for functional gene analysis during T lymphocyte activation and differentiation in vitro as well as in vivo.Key words: Gene expression . Immune regulation . T cells Introduction RNA interference (RNAi) is an evolutionarily conserved process by which double-stranded small interfering RNA (siRNA) induces sequence-specific, post-transcriptional gene silencing [1][2][3]. Given its ease of application, its high efficiency and remarkable specificity, RNAi holds great promise for broad in vitro and in vivo application in all biomedical areas. Most importantly, RNAi is directly applicable to essentially all somatic cell types including human primary cells, which makes it an invaluable tool to study human gene function and may enable new therapeutic approaches [4,5].Within the immune system T lymphocytes are one major target for siRNA-based gene silencing, since they are key regulators of immune responses and involved in many immune related disorders, like autoimmunity, chronic inflammation or lymphoma.Despite this high potential, the lack of protocols for efficient and sustained siRNA delivery into primary mammalian cells is currently the major obstacle to the use of RNAi. In particular, primary lymphocytes are highly resistant to non-viral transfection using cationic lipids and polymer reagents [6][7][8]. Although lymphocytes can be transfected by electroporation in vitro, this method so far has been rather inefficient, limited to activated cells and was complicated by a severe impairment of cell function and cell viability [9,10]. Recently, a promising approach for the 2616in vivo targeting of lymphocytes has been reported using antibody-protamine fusion proteins to deliver siRNA [11]. However, recent data from small-hairpin RNA transgenic mice indicate that in T lymphocytes the RNAi machinery itself works inefficiently as compared with other cell types [12]. In fact, due to the aforementioned problems with siRNA delivery, the parameters determining RNAi efficien...

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“…Second, using conditional Gata3 knockout mice, it was observed that deletion in CD4/CD8 double-negative thymocytes blocked T cell development at the DN3 stage [42]. Third, deleting npg cells, although development of CD8 single positive T cells appears normal [42,43]. We prepared a conditional Gata3 knockout by flanking exon 4 of the Gata3 gene with two loxP sites [43].…”

Section: The Central Role Of Gata-3 In Reinforcing Th2 Responsesmentioning

confidence: 99%

GATA-3 promotes Th2 responses through three different mechanisms: induction of Th2 cytokine production, selective growth of Th2 cells and inhibition of Th1 cell-specific factors

et al. 2006

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Naïve CD4 T cells can differentiate into at least two different types of T helpers, Th1 and Th2 cells. Th2 cells, capable of producing IL-4, IL-5 and IL-13, are involved in humoral immunity against extracellular pathogens and in the induction of asthma and other allergic diseases. In this review, we summarize recent reports regarding the transcription factors involved in Th2 differentiation and cell expansion, including Stat5, Gfi-1 and GATA-3. Stat5 activation is necessary and sufficient for IL-2-mediated function in Th2 differentiation. Enhanced Stat5 signaling induces Th2 differentiation independent of IL-4 signaling; although it does not up-regulate GATA-3 expression, it does require the presence of GATA-3 for its action. Gfi-1, induced by IL-4, promotes the expansion of GATA-3-expressing cells. Analysis of conditional Gata3 knockout mice confirmed the critical role of GATA-3 in Th2 cell differentiation (both IL-4 dependent and IL-4 independent) and in Th2 cell proliferation and also showed the importance of basal GATA-3 expression in inhibiting Th1 differentiation.

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Conditional deletion of Gata3 shows its essential function in TH1-TH2 responses

Cited by 564 publications

References 48 publications

IL‐18 stimulates IL‐13‐mediated IFN‐γ‐sensitive host resistance in vivo

IL‐18 stimulates IL‐13‐mediated IFN‐γ‐sensitive host resistance in vivo

siRNA stabilization prolongs gene knockdown in primary T lymphocytes

GATA-3 promotes Th2 responses through three different mechanisms: induction of Th2 cytokine production, selective growth of Th2 cells and inhibition of Th1 cell-specific factors

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