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  Director : Sandra Pellegrini (pellegri@pasteur.fr)


  abstract

 

Type I interferons (IFN-a/b) are a family of innate cytokines that also actively participate to the shaping of adaptive immune T helper 1 responses. Present work in my laboratory focuses on different aspects of the biology of IFN-a/b, from the analysis of the receptor/Jak kinase complexes and novel Jak interactors, to the study of the role that these cytokines may play in the development of human T lymphocytes.



  report

cale

In addition to their potent antiviral activity, the a IFNs and IFN-b are now recognized as being front line players at the interface between innate and adaptive immune responses against pathogens. Our work focuses on molecular aspects of the cellular responses to IFN-a/b in the human system. We have progressed in the analysis of the receptor/kinase complex and of novel components of this signaling pathway. Moreover, we are addressing questions related to the role of IFN-a/b in differentiation and survival of helper T lymphocytes.

Our research program develops around three major axes. From our original finding of the role of the tyrosine kinase Tyk2 in stabilizing the IFNAR1 receptor chain at the cell surface, we are presently interested in defining the role of the tyrosine kinases Tyk2 and Jak1 in receptor trafficking. In fact, little is known of the molecular mechanisms that regulate biosynthesis, assembly and spatio-temporal dynamics of the receptor/Jak complexes. A second axis concerns the biochemical and functional characterization of two unrelated Tyk2-interacting proteins, identified in a two-hybrid screen. A third aspect of our work concerns IFN-a/b and its functional impact on human T lymphocytes. We have established a model system of naïve CD4 T cells purified from cord blood that can be stimulated in vitro to develop along different developmental programs. In this system we have uncovered a different reactivity to IFN-a/b of naïve uncommitted cells versus activated cells and we plan to define the molecular mechanisms underlying these changes.

The tyrosine kinase Tyk2 and trafficking of IFNAR1 J. Ragimbeau, E. Dondi in collaboration with A. Alcover (U. of the Biologie of Cellular Interactions, I. P.) , P. Eid (CNRS, Villejuif) and G. Uzé (CNRS, Montpellier)

We have demonstrated that Tyk2 augments the level of the IFN-a/b receptor chain IFNAR1 at the plasma membrane and to do so Tyk2 down-regulate its internalisation rate. Based on our model, Tyk2, by associating to IFNAR1 through its FERM domain, masks motifs required for receptor internalisation. This work, along with recent data from other laboratories, demonstrate that Jak enzymes act not solely as key effectors of cytokine receptors, but also upregulate, through different mechanisms, receptor surface levels. On these premises, we are now pursuing the study of the role of Jaks in ligand-induced endocytosis and trafficking of the receptor.

Study of two novel interactors of the tyrosine kinase Tyk2, identified by two hybrid C. Steindler and Z. Li

The amino-terminal moiety of the Jak proteins carries the molecular determinants for binding to cytokine receptors. Sequence analysis of the amino-terminal region of JAKs has revealed the presence of a band 4.1-related domain or FERM. This domain is present in cytoskeletal proteins of the ERM family (ezrin, radixin, moesin) where it mediates intramolecular interactions and membrane targeting. A yeast two-hybrid screen was performed using as bait the FERM domain of Tyk2. Functional and biochemical characterization of two partners is ongoing. We have progressed in the characterization of Pot2, which belongs to a family of 3 genes. Pot2 and Pot2A transcritps are strongly expressed in the neural system and weakly in hematopoietic cell lineages. These proteins rich in coiled-coils associate through their N-terminal region to the microtubule cytoskeleton. We are studying the function of Pot2 in two cellular systems, the Jurkat T cell line and the rat pheochromocytoma PC12 cell line. The second interactor, Pot-1, is a novel protein expressed in most tissues which appears localized a vesicular compartment. To down-regulate Pot1 expression and study its function, RNA interference will be used.

L'IFN-a/b et les lymphocytes T humains E. Dondi, in collaboration with L. Rogge (Lab. of Immunoregulation, I. P.), G. Lutfalla and G. Uzé (CNRS, Montpellier)

A critical aspect of the immune response to pathogens is mediated by the CD4 and CD8 T lymphocytes which, upon suitable activation, differentiate and expand into functionally distinct subsets. IFN-a/b is produced in response to a viral or bacterial infection and performs multiple immunoregulatory functions, instructing naïve T cells to develop down the Th1 developmental pathway. We have studied the response to IFN-a/b of primary naïve T lymphocytes purified from cord blood and differentiated in vitro into effector Th1/Th2 cells. Our analysis shows that stimulation of naïve lymphocytes, through TCR engagement, leads to a change in reactivity to IFN-a/b, measured as proliferative potential and target gene induction. Briefly, cell cycle entry of naïve cells is retarded by IFN treatment, while the ongoing proliferation of pre-activated lymphocytes is unaffected. This different reactivity may allow rapid expansion of effector cells in a context of viral infections where large quantity of this anti-proliferative cytokine is produced in secondary lymphoid organs. Moreover, the induction of target genes is 10 to 20 fold reduced in effector lymphocytes as compared to their naïve precursors. This down-modulation takes place at the time of T cell activation and is part of the complex genetic program of T cell differentiation. We are currently exploring the molecular mechanisms underlying these changes and we are also interested in studying the impact of IFN-a/b on the generation of memory T cells.

IFN-a/b gene induction in human dendritic cells infected by Mycobacterium tuberculosis E. Dondi in collaboration with E. Coccia (Istituto Superirore di Sanità, Rome, Italy)

We have participated in the analysis of the indution profile of IFN-a/b genes in a model system of DC infected Mycobacterium tuberculosis (Mtb). The infection induces an early secretion of IFN-b and at a later stage secretion of a single subtype of IFN-a/b (a1). The signaling steps of this sequential induction have been dissected by monitoring the transcription factors activated during bacterial infection (NFkB, IRF-3, Stat1/2, IRF-7). Our results suggest the existence of a cascade of events analogous that the one that occurs upon viral infection.

Keywords: interferon, cytokine, receptor, signaling, lymphocyte, immunologie, cell biologie, biochemistry



  publications

puce Publications of the unit on Pasteur's references database


  personnel

  Office staff Researchers Scientific trainees Other personnel
  Houssin Wendy, Wendy.Houssin@pasteur.fr Dondi Elisabetta, Post-Doc,edondi@pasteur.fr

Steindler Corinna, Post-Doc,costei@pasteur.fr

Li Zhi, Post-Doc,zhili@pasteur.fr

Ragimbeau Josiane, Engineer,jragimbe@pasteur.fr

Activity Reports 2002 - Institut Pasteur
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