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  Director : THEZE Jacques (jtheze@pasteur.fr)


  abstract

 

IL-2 is a major cytokine of the immune system. It interacts with a receptor (RIL-2) made up of three sub-units (a, b and g). The work we are conducting in this field is focused on three main research areas: 1) the role played by IL-2 in controlling the amplitude of T cell responses and the maintenance of homeostasis, 2) the characterization of IL-2 signaling routes through a study of human IL-2 mimetic peptide, 3) the effect of IL-2/RIL-2 system deregulation on the immune deficiency induced during HIV infection.



  report

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IL-2 mechanism of action: control of the equilibrium between activation/differentiation and maintenance of homeostasis. (P. Chastagner, J. Reddy)

We have sought to identify new genes whose expression is under the control of IL-2. A subtractive CDNA approach was employed for a cell line able to grow in IL-2 or IL-4. From this we were able to select 66 different transcripts. The expression kinetics of 8 of these transcripts suggested that they are induced during activation/differentiation phenomena triggered by IL-2 in T lymphocytes. The first group of these genes code for factors involved in transcription control e.g. CCCTC-binding factor, Jun inhibitor factor-1 and transcription factors E2F-4 and AMP-responsive element-binding protein, zhx-1. IL-2 also increases the expression of atubulin, a cytoskeletal protein, and multifunctional proteins such as b-catenin and nucleolin. In addition, we demonstrated that TNF-b is one of the first genes to be induced by IL-2 through a mechanism identified previously in the induction of the IL-2Ra chain (activation of the Jak/STAT pathway and presence of a binding site for STAT5 upstream of the TNF-bgene). The study of the expression of these genes was continued in vivo using mice where the IL-2 gene is either disrupted or not (IL-2-/-, IL-2-/+). IL-2-/- mice are characterized in terms of secondary lymphoid organs by increased proliferation of T lymphocytes but a thymus that is normal both in structure and function. Our studies in IL-2 deficient mice showed that some of the genes coding for secondary lymphoid organs were not expressed. As far as these organs are concerned, other genes of the TNF group (TNF-a, LT-b, TNFR1 and TNFR2) have been shown — in the same manner as TNF-bto be under the control of IL-2. The critical role played by IL-2 in the expression of these genes suggests that they may be involved in controlling the activation and maintenance of homeostasis in peripheral T cells. Recently, we showed that the proliferation and lesser susceptibility to opoptois of T cells taken from IL-2-/- mice is associated with enhanced expression of cFLIP, a protein that inhibits the activation of TNF-R- or FAS-dependent apoptosis.

Characterization of a new human IL-2Rb agonist. (R. Eckenberg,, J-L. Moreau, F. Gesbert).

In the framework of structure-function studies of human IL-2, we characterized a new agonist of the RIL-2 b chain known as peptide p1-30. This peptide consists of the first 30 amino acids of IL-2. A study of its physicochemical properties in solution showed that the molecule folds to form an a helix. Several of these molecules group together to form tetramers. Modeling studies suggest that these tetramers associate with the dimers of the human IL-2R b chain. In vitro, p1-30 induces similar biological activities to those of a growth factor or survival factor in cell lines expressing certain combinations of the three chains of human IL-2R. We plan to study the signaling routes activated by the IL-2Rb2 dimers and investigate more deeply the mechanisms involved in IL-2 transduction signals via the bg, abg and b2 heterodimers. Synergy was observed with IL-2, IL-4, IL-9 and IL-15, i.e. all cytokines which include the g c chain in the composition of their receptor. We undertook to determine the potential cell targets of p1-30 among the lymphocytes in the bloodstream. LAK activity was induced and IFN-gwas secreted. At the same time, we noted that cells of the NK and CD8low subpopulations — constitutively expressing the IL-2 b chain — showed an activated cell phenotype. The studies conducted on p1-30 are of fundamental value since they provide a more precise description of the function and organization of the IL-2 receptor. These studies also open the way toward future therapeutic applications. Results obtained in our laboratory and by others indicate that p1-30 or p1-30-derived molecules can activate cytotoxic cells with a higher therapeutic index than IL-2. In our continuation along this avenue of research, we have conducted studies in a transgenic mouse model that expresses the b chain of human IL-2R.

Involvement of IL-2 and its receptor in the immune disease associated with HIV infection and HIV immune therapy. (D. David, H. Keller, L. Bani, S. Beq, V. Pasquier, M. Kryworuchko, J-H. Colle.)

Most of the lymphocytes in the bloodstream are devoted to meeting new antigenic challenges. Any reduction in the size or alteration in the functional properties of this cell contingent has an impact on resistance to infectious agents.

We analyzed the expression of the three IL-2R chains on the surface of lymphocytes isolated from the blood of HIV+ patients, and compared this with expression in normal individuals. Two expression profiles were observed in the normal individuals, corresponding to the separation of cells into those involved in non-specific and specific responses. The NK cells and monocytes showed abundant expression of the b and g chains respectively. Most T lymphocytes (CD4, CD8) and B lymphocytes did not show any surface expression of the IL-2R chains. However, g chain was found in an intracellular compartment of the CD4, CD8, NK and B lymphocytes. The HIV+ patients with an elevated viral load showed surface expression of the three RIL-2 chains on a large proportion of CD8 and B lymphocytes. The B lymphocytes in these patients showed a response to IL-2, contrasting with insensitivity of their CD8 and a reduction in the NK response to this cytokine. We also studied the status of the IL-2/IL-2R complex in patients receiving HAART and showing reduced viral load (virus particles below the limit of detection). Thanks to HAART, most patients show a significant increase in the peripheral CD4 count. This is accompanied by restored CD8 and CD4 response to IL-2. However, the small proportion of patients whose CD4 count is not increased by HAART continues to show a defective response to IL-2. The impaired capacity of CD4 from HIV+ patients to produce IL-2 has for a long time been considered to be a critical factor in the mechanism of immunodeficiency. The IL-2R functional anomalies we have demonstrated in the different patients groups also contribute to the immune deficiency of HIV+ patients. More recently, we analyzed the IL-2/IL-2R complex in patients who were given IL-2 because HAART had little effect on their CD4 counts. We noted a rapid rise in the CD4 count associated with the following changes in the CD4 cells: a decrease in spontaneous susceptibility to cell death by apoptosis, an increase in Bcl-2 expression and the acquisition of a response to IL-2. The results of these studies clearly illustrate the numerous facets of HIV-induced immune deficiency. Our current studies aim to describe the nature of the defect in the response to IL-2R, determine its relationship with immune deficiency and identify the mechanism by which immunotherapy restores the CD4 count in the peripheral compartment.



  publications

puce Publications of the unit on Pasteur's references database


  personnel

  Office staff Researchers Scientific trainees Other personnel
 

BREGEAT Annick. émail : abregeat@pasteur.fr

COLLE Jean-Hervé. Pasteur. émail : jhcolle@pasteur.fr

GESBERT Franck. Pasteur. émail : fgesbert@pasteur.fr

BEQ Stéphanie. PhD student é.mail : sbeq@pasteur.fr

KRYWORUCHKO Marko. Postdoc. é.mail : mkrywor@pasteur.fr

PASQUIER Virginie PhD student émail : vmazard@pasteur.fr

Engineer : MOREAU Jean-Louis. émail : jlmoreau@pasteur.fr

Technician : CHASTAGNER Patricia. eémail : pchasta@pasteur.fr


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