IL-12, but not IFN-α, promotes STAT4 activation and Th1 development in murine CD4 T cells expressing a chimeric murine/human Stat2 gene

Center for Immunology and Department of Molecular Biology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390, USA.
The Journal of Immunology (Impact Factor: 4.92). 02/2005; 174(1):294-301. DOI: 10.4049/jimmunol.174.1.294
Source: PubMed

ABSTRACT Humans and mice have evolved distinct pathways for Th1 cell development. Although IL-12 promotes CD4(+) Th1 development in both murine and human T cells, IFN-alphabeta drives Th1 development only in human cells. This IFN-alphabeta-dependent pathway is not conserved in the mouse species due in part to a specific mutation within murine Stat2. Restoration of this pathway in murine T cells would provide the opportunity to more closely model specific human disease states that rely on CD4(+) T cell responses to IFN-alphabeta. To this end, the C terminus of murine Stat2, harboring the mutation, was replaced with the corresponding human Stat2 sequence by a knockin targeting strategy within murine embryonic stem cells. Chimeric m/h Stat2 knockin mice were healthy, bred normally, and exhibited a normal lymphoid compartment. Furthermore, the murine/human STAT2 protein was expressed in murine CD4(+) T cells and was activated by murine IFN-alpha signaling. However, the murine/human STAT2 protein was insufficient to restore full IFN-alpha-driven Th1 development as defined by IFN-gamma expression. Furthermore, IL-12, but not IFN-alpha, promoted acute IFN-gamma secretion in collaboration with IL-18 stimulation in both CD4(+) and CD8(+) T cells. The inability of T cells to commit to Th1 development correlated with the lack of STAT4 phosphorylation in response to IFN-alpha. This finding suggests that, although the C terminus of human STAT2 is required for STAT4 recruitment and activation by the human type I IFNAR (IFN-alphabetaR), it is not sufficient to restore this process through the murine IFNAR complex.

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Available from: David Farrar, Sep 28, 2015
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    • "The insertion in mouse STAT2 alters the C-terminus and selectively disrupts its capacity to activate STAT4 [27]. However, this notion was negated by the finding that mice expressing a chimeric mouse STAT2 with human TAD domain were unable to activate STAT4 and trigger Th1 differentiation [28]. In fact, overexpression of mouse STAT2 in human U6 cell line that does not express STAT2 rescues IFNα response [21]. "
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    ABSTRACT: Interferons (IFNs) are key regulators for both innate and adaptive immune responses. By screening ENU-mutagenized mice, we identified a pedigree- P117 which displayed impaired response to type I, but not type II, IFNs. Through inheritance test, genetic mapping and sequencing, we found a T to A point mutation in the 5' splice site of STAT2 intron 4-5, leading to cryptic splicing and frame shifting. As a result, the expression of STAT2 protein was greatly diminished in the mutant mice. Nonetheless, a trace amount of functional STAT2 protein was still detectable and was capable of inducing, though to a lesser extent, IFNalpha-downstream gene expressions, suggesting that P117 is a STAT2 hypomorphic mutant. The restoration of mouse or human STAT2 gene in P117 MEFs rescued the response to IFNalpha, suggesting that the mutation in STAT2 is most likely the cause of the phenotypes seen in the pedigree. Development of different subsets of lymphocytes appeared to be normal in the mutant mice except that the percentage and basal expression of CD86 in splenic pDC and cDC were reduced. In addition, in vitro Flt3L-dependent DC development and TLR ligand-mediated DC differentiation were also defective in mutant cells. These results suggest that STAT2 positively regulates DC development and differentiation. Interestingly, a severe impairment of antiviral state and increased susceptibility to EMCV infection were observed in the mutant MEFs and mice, respectively, suggesting that the remaining STAT2 is not sufficient to confer antiviral response. In sum, the new allele of STAT2 mutant reported here reveals a role of STAT2 for DC development and a threshold requirement for full functions of type I IFNs.
    Journal of Biomedical Science 02/2009; 16(1):22. DOI:10.1186/1423-0127-16-22 · 2.76 Impact Factor
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    • "Therefore, although the precise role of T-bet in autoimmune inflammation remains elusive, these data show clearly that T-bet is not required for IL-17 production from CD4 T cells, reiterating the notion that Th17 effector cells develop independently of Th1 and Th2 cells. Consistent with this, STAT4-deficient mice are resistant to the chronic inflammatory disorders in which the IL-23–IL-17 pathway drives autoimmune disease, EAE, suggesting that STAT4 is required for development of pathogenic Th17 cells (Persky et al., 2005). Indeed, signaling through the IL-23R complex has been shown to phosphorylate STAT1, STAT3, and STAT4, as well as to induce the formation of STAT3/STAT4 heterodimers, suggesting that both STAT3 and STAT4 might be important for IL-17 production from CD4 T cells (Lin et al., 2005). "
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    ABSTRACT: The T help 1 (Th1) and Th2 cell classification have provided the framework for understanding CD4(+) T cell biology and the interplay between innate and adaptive immunity for almost two decades. Recent studies have defined a previously unknown arm of the CD4(+) T cell effector response, the Th17 lineage, which promises to change our understanding of immune regulation, immune pathogenesis and host defense. The factors that specify differentiation of IL-17 producing effector T cells from naïve T cell precursors are being rapidly discovered and are providing insights into mechanisms by which signals from cells of the innate immune system guide alternative pathways of Th1, Th2, or Th17 development. In this review, we will focus on recent studies that have identified new subsets of Th cells, new insights regarding the induced generation and differentiation mechanisms of Th17 cells and immune regulatory effects.
    Journal of Cellular Physiology 05/2007; 211(2):273-8. DOI:10.1002/jcp.20973 · 3.84 Impact Factor
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    • "Our previous studies demonstrated that although the human STAT2 C-terminus was required to promote IFN-α-dependent STAT4 activation in human fibroblasts (Farrar et al., 2000a), it was not sufficient to restore this pathway when expressed in murine T cells (Persky et al., 2005). This finding predicts that additional species-specific components of the IFNAR complex regulate STAT4 tyrosine phosphorylation. "
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    ABSTRACT: CD4(+) T cells regulate adaptive responses to pathogens by secreting unique subsets of cytokines that mediate inflammatory processes. The innate cytokines IL-12 and IFN-alpha/beta regulate type I responses and promote acute IFN-gamma secretion through the activation of the STAT4 transcription factor. Although IL-12-induced STAT4 activation is a conserved pathway across species, IFN-alpha/beta-dependent STAT4 phosphorylation does not occur as efficiently in mice as it does in human T cells. In order to understand this species-specific pathway for IFN-alpha/beta-dependent STAT4 activation, we have examined the molecular basis of STAT4 recruitment by the human IFNAR. In this report, we demonstrate that the N-domain of STAT4 interacts with the cytoplasmic domain of the human, but not the murine IFNAR2 subunit. This interaction mapped to a membrane-proximal segment of the hIFNAR2 spanning amino acids 299-333. Deletion of this region within the hIFNAR2 completely abolishes IFN-alpha/beta-dependent STAT4 tyrosine phosphorylation when expressed in human IFNAR2-deficient fibroblasts. Thus, the human IFNAR2 cytoplasmic domain serves to link STAT4 to the IFNAR as a pre-assembled complex that facilitates cytokine-driven STAT4 activation.
    Molecular Immunology 04/2007; 44(8):1864-72. DOI:10.1016/j.molimm.2006.10.006 · 2.97 Impact Factor
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