Molecular interactions determining defective Th2 cell differentiation of IRF4 deficient T helper cells | |
| Kennziffer | 20.261.15p |
| Projektleiter/in | Prof. Dr. Michael Lohoff |
| Weitere Bearbeiter/innen | Anne Brüstle |
| Kurzbeschreibung |
Background Almost 20 years ago, T helper (h) cells were subdivided into two subsets, Th1 and Th2, based on the cytokines they produce. The existence of cytokine patterns of either Th1 or Th2 type was later described in many diseases in vivo, such as infections with all types of pathogens, autoimmune diseases including multiple sclerosis and even cancer (1). Importantly, in a given disease either Th1 or Th2 cytokine patterns can exist and the emerging Th type may decide the outcome of the disease. We have established two infectious mouse models, in which the developing Th phenotype is of prime importance for the course of disease. After infection with the protozoan parasite Leishmania major, resistant mouse strains clear the infection due to expansion of Th1 cells which, by producing Interferon (IFN)-g, activate macrophages to kill this intracellular pathogen. In contrast, infected susceptible mice die after Th2 cell expansion which deactivate macrophages by secreting IL-4 and IL-10 (1). Infection of mice with the bacterium Helicobacter pylori causes gastritis and gastric atrophy only in mice, that expand Th1 cytokines, while Th2 cells seem to protect from gastritis (2). Th1 and Th2 cells differentiate from common precursors and the presence of IL-12, acting through STAT4, favours Th1 cell differentiation, while IL-4 uses STAT6 to mediate Th2 cell differentiation. However, alternative pathways seem to exist. Two transcription factors are absolutely essential, namely T-bet for Th1 and GATA3 (which induces its own expression) for Th2 cell differentiation (1). Our lab has contributed to this topic by defining a role of transciption factors of the IRF family for Th differentiation (3-5). Among other activities, IRF1 and IRF2 bind to common response elements in the promoters of the IL-4 and IL-12 genes to influence IL-4 and IL-12 production. In addition, IRF4, which is expressed in cells of the hematopoietic lineage, regulates Th2 differentiation by influencing transcription of GATA3. However, how IRF4 and GATA3 are connected is still unknown. A trivial possibility is that IRF4 binds to the GATA3 promoter, but so far, no such binding has been described. Many known IRF4 binding partners (IRF1 and IRF2, NFAT factors, STAT6, BCL-6, PU.1) which considerably influence the ativity of IRF4, further complicate the issue (6-9). A very attractive candidate that might influence Th2 cell differentiation is PU.1 and in the last 15 months of the "Graduiertenkolleg" we have established that PU.1 is expressed in Th2, but not Th1 cells. Thus, a dimer of IRF4 and PU.1 might act in Th2, but not Th1 cells. Aims a) Dr Hendrik (Immunologie, Rotterdam) has established a knock-in mouse in which GATA-3 is replaced by a lacZ reporter insertion. As one possibility to address the link between IRF4 and GATA-3, these mice will be crossed with IRF4-deficient mice. Here, one can study, if IRF4 deficiency causes similar GATA-3 deficiency in several cell types or whether the defect is restricted to T helper cells. b) There are established interactions between CBP/p300 and IRF1 as well as between CBP/p300 and GATA-1. Also, a B cell specific inactivation of CBP/p300 leads to a phenotype (splenomegaly, lack of mature B cells) similar to the one of IRF4 deficient mice. In order to search for a potential molecular interaction between CBP/p300 and IRF4 (in collaboration with Werner Lutz, IMT, Marburg), such an interaction will be first investigated in vitro. Thereafter, the mice with the "floxed" CBP/p300 gene will be used to create T cell specific deficiency of CBP/p300 and to analyse the T helper cell phenotype in such animals. c) Using siRNA analysis, it will be investigated whether a deficiency in PU.1 creates a T cell defect similar to the one observed in IRF4 -/- Th cells. d) Microarray analyses will be used to identify pathways which are dysregulated in IRF4 -/- Th cells during Th cell differentiation. e) IRF4 interacts with PU.1, which in turn interacts with GATA-1. Therefore, in a joint analysis with Dr Philipsen from Rotterdam, it will be investigated whether a deficiency in IRF4 has implications for the expression of other GATA members than just GATA-3. This analysis will be made in different cell types of the hematopoietic lineage. In parallel, it will be tested whether GATA-1 is expressed at some stage during Th1 or Th2 cell differentiation. If so, a mouse carrying a "floxed"GATA-1 gene will be used to create a T cell specific GATA-1 defect. It will be studied whether such a mouse has a phenotype with similarities to the IRF4-deficient mouse. In addition, a genetic intercross between GATA-1 and IRF4-deficient mice will establish, whether part of the phenotype created by IRF4-deficiency can be explained by dysregulated GATA-1 activity.
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| Laufzeit | 01.01.2002 - 31.12.2010 |
| Finanzierung | DFG |
| Zugehörigkeit | Transcriptional control in developmental processes Internationales Graduiertenkolleg Universität Marburg, Universität Gießen, Universität Rotterdam (DFG-GRK 767) |
