N Mathieu

French Institute of Health and Medical Research, Lutetia Parisorum, Île-de-France, France

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Publications (9)69.53 Total impact

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    ABSTRACT: To assess the role of the T cell receptor (TCR) beta gene enhancer (Ebeta) in regulating the processing of VDJ recombinase-generated coding ends, we assayed TCRbeta rearrangement of Ebeta-deleted (DeltaEbeta) thymocytes in which cell death is inhibited via expression of a Bcl-2 transgene. Compared with DeltaEbeta, DeltaEbeta Bcl-2 thymocytes show a small accumulation of TCRbeta standard recombination products, including coding ends, that involves the proximal Dbeta-Jbeta and Vbeta14 loci but not the distal 5' Vbeta genes. These effects are detectable in double negative pro-T cells, predominate in double positive pre-T cells, and correlate with regional changes in chromosomal structure during double negative-to-double positive differentiation. We propose that Ebeta, by driving long range nucleoprotein interactions and the control of locus expression and chromatin structure, indirectly contributes to the stabilization of coding ends within the recombination processing complexes. The results also illustrate Ebeta-dependent and -independent changes in chromosomal structure, suggesting distinct modes of regulation of TCRbeta allelic exclusion depending on the position within the locus.
    Journal of Biological Chemistry 06/2003; 278(20):18101-9. DOI:10.1074/jbc.M212647200 · 4.60 Impact Factor
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    ABSTRACT: We demonstrate that overexpression of Pim-1, a cytoplasmic serine/threonine kinase of poorly defined function, results in the development of substantial numbers of CD4(+)CD8(+) double-positive thymocytes in two independent knock-out mouse models (i.e. the RAG-1-deficient and TCRbeta gene enhancer-deleted mice) in which production of a functionally rearranged TCRbeta gene (hence the pre-TCR) is impaired. This activity of Pim-1, however, does not affect signaling through the Ras/Raf/MAP kinase cascade nor signaling which mediates suppression of TCRbeta gene recombination (i.e. allelic exclusion). While overexpression of Pim-1 positively affects cell cycle progression in selected CD4(-)CD8(-) double-negative precursors, it did not affect expression of components of the cell cycle machinery, with the exception of the G(1)-specific phosphatase Cdc25A upon antigen receptor stimulation. We propose that Pim-1 acts downstream, or in parallel, to pre-TCR-mediated selection as one factor involved in the proliferative expansion of beta-selected pre-T cells.
    International Immunology 11/2000; 12(10):1389-96. DOI:10.1093/intimm/12.10.1389 · 3.18 Impact Factor
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    ABSTRACT: Gene targeting studies have shown that T cell receptor (TCR)-beta gene expression and recombination are inhibited after deletion of an enhancer (Ebeta) located at the 3' end of the approximately 500-kb TCR-beta locus. Using knockout mouse models, we have measured, at different regions throughout the TCR-beta locus, the effects of Ebeta deletion on molecular parameters believed to reflect epigenetic changes associated with the control of gene activation, including restriction endonuclease access to chromosomal DNA, germline transcription, DNA methylation, and histone H3 acetylation. Our results demonstrate that, in early developing thymocytes, Ebeta contributes to major chromatin remodeling directed to an approximately 25-kb upstream domain comprised of the Dbeta-Jbeta locus regions. Accordingly, treatment of Ebeta-deleted thymocytes with the histone deacetylase inhibitor trichostatin A relieved the block in TCR-beta gene expression and promoted recombination within the Dbeta-Jbeta loci. Unexpectedly, however, epigenetic processes at distal Vbeta genes on the 5' side of the locus and at the 3' proximal Vbeta14 gene appear to be less dependent on Ebeta, suggesting that Ebeta activity is confined to a discrete region of the TCR-beta locus. These findings have implications with respect to the developmental control of TCR-beta gene recombination, and the process of allelic exclusion at this locus.
    Journal of Experimental Medicine 10/2000; 192(5):625-36. · 13.91 Impact Factor
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    ABSTRACT: Gene targeting studies have shown that T cell receptor (TCR)-β gene expression and recombination are inhibited after deletion of an enhancer (Eβ) located at the 3′ end of the ∼500-kb TCR-β locus. Using knockout mouse models, we have measured, at different regions throughout the TCR-β locus, the effects of Eβ deletion on molecular parameters believed to reflect epigenetic changes associated with the control of gene activation, including restriction endonuclease access to chromosomal DNA, germline transcription, DNA methylation, and histone H3 acetylation. Our results demonstrate that, in early developing thymocytes, Eβ contributes to major chromatin remodeling directed to an ∼25-kb upstream domain comprised of the Dβ-Jβ locus regions. Accordingly, treatment of Eβ-deleted thymocytes with the histone deacetylase inhibitor trichostatin A relieved the block in TCR-β gene expression and promoted recombination within the Dβ-Jβ loci. Unexpectedly, however, epigenetic processes at distal Vβ genes on the 5′ side of the locus and at the 3′ proximal Vβ14 gene appear to be less dependent on Eβ, suggesting that Eβ activity is confined to a discrete region of the TCR-β locus. These findings have implications with respect to the developmental control of TCR-β gene recombination, and the process of allelic exclusion at this locus.
    Journal of Experimental Medicine 09/2000; 192(5):625-636. DOI:10.1084/jem.192.5.625 · 13.91 Impact Factor
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    ABSTRACT: T cell differentiation in the mouse thymus is an intricate, highly coordinated process that requires the assembly of TCR complexes from individual components, including those produced by the precisely timed V(D)J recombination of TCR genes. Mice carrying a homozygous deletion of the TCR beta transcriptional enhancer (E beta) demonstrate an inhibition of V(D)J recombination at the targeted TCR beta locus and a block in alpha beta T cell differentiation. In this study, we have characterized the T cell developmental defects resulting from the E beta-/- mutation, in light of previously reported results of the analyses of TCR beta-deficient (TCR beta-/-) mice. Similar to the latter mice, production of TCR beta-chains is abolished in the E beta-/- animals, and under these conditions differentiation into cell-surface TCR-, CD4+CD8+ double positive (DP) thymocytes depends essentially on the cell-autonomous expression of TCR delta-chains and, most likely, TCR gamma-chains. However, contrary to previous reports using TCR beta-/- mice, a minor population of TCR gamma delta+ DP thymocytes was found within the E beta-/- thymi, which differ in terms of T cell-specific gene expression and V(D)J recombinase activity, from the majority of TCR-, alpha beta lineage-committed DP thymocytes. We discuss these data with respect to the functional role of E beta in driving alpha beta T cell differentiation and the mechanism of alpha beta T lineage commitment.
    The Journal of Immunology 09/2000; 165(3):1364-73. DOI:10.4049/jimmunol.165.3.1364 · 5.36 Impact Factor
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    ABSTRACT: T cell differentiation in the mouse thymus is an intricate, highly coordinated process that requires the assembly of TCR complexes from individual components, including those produced by the precisely timed V(D)J recombination of TCR genes. Mice carrying a homozygous deletion of the TCRβ transcriptional enhancer (Eβ) demonstrate an inhibition of V(D)J recombination at the targeted TCRβ locus and a block in αβ T cell differentiation. In this study, we have characterized the T cell developmental defects resulting from the Eβ−/− mutation, in light of previously reported results of the analyses of TCRβ-deficient (TCRβ−/−) mice. Similar to the latter mice, production of TCRβ-chains is abolished in the Eβ−/− animals, and under these conditions differentiation into cell-surface TCR−, CD4+CD8+ double positive (DP) thymocytes depends essentially on the cell-autonomous expression of TCRδ-chains and, most likely, TCRγ-chains. However, contrary to previous reports using TCRβ−/− mice, a minor population of TCR γδ+ DP thymocytes was found within the Eβ−/− thymi, which differ in terms of T cell-specific gene expression and V(D)J recombinase activity, from the majority of TCR−, αβ lineage-committed DP thymocytes. We discuss these data with respect to the functional role of Eβ in driving αβ T cell differentiation and the mechanism of αβ T lineage commitment.
    The Journal of Immunology 08/2000; 165(3):1364-1373. · 5.36 Impact Factor
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    ABSTRACT: V(D)J recombination in differentiating lymphocytes is a highly regulated process in terms of both cell lineage and the stage of cell development. Transgenic and knockout mouse studies have demonstrated that transcriptional enhancers from antigen receptor genes play an important role in this regulation by activating cis-recombination events. A striking example is the T-cell receptor beta-chain (TCRbeta) gene enhancer (Ebeta), which in the mouse consists of at least seven nuclear factor binding motifs (betaE1 to betaE7). Here, using a well-characterized transgenic recombination substrate approach, we define the sequences within Ebeta required for recombination enhancer activity. The Ebeta core is comprised of a limited set of motifs (betaE3 and betaE4) and an additional previously uncharacterized 20-bp sequence 3' of the betaE4 motif. This core element confers cell lineage- and stage-specific recombination within the transgenic substrates, although it cannot bypass the suppressive effects resulting from transgene integration in heterochromatic centromeres. Strikingly, the core enhancer is heavily occupied by nuclear factors in immature thymocytes, as shown by in vivo footprinting analyses. A larger enhancer fragment including the betaE1 through betaE4 motifs but not the 3' sequences, although active in inducing germ line transcription within the transgenic array, did not retain the Ebeta recombinational activity. Our results emphasize the multifunctionality of the TCRbeta enhancer and shed some light on the molecular mechanisms by which transcriptional enhancers and associated nuclear factors may impact on cis recombination, gene expression, and lymphoid cell differentiation.
    Molecular and Cellular Biology 02/2000; 20(1):42-53. DOI:10.1128/MCB.20.1.42-53.2000 · 5.04 Impact Factor
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    ABSTRACT: Deletion of the TCRbeta transcriptional enhancer (Ebeta) results in nearly complete inhibition of V(D)J recombination at the TCRbeta locus and a block in alpha beta T cell development. This result, along with previous work from many laboratories, has led to the hypothesis that transcriptional enhancers affect V(D)J recombination by regulating the accessibility of the locus to the recombinase. Here we test this hypothesis by performing a detailed analysis of the recombination defect in Ebeta-deleted (Ebeta-/-) mice using assays that detect various reaction intermediates and products. We found double-strand DNA breaks at recombination signal sequences flanking Dbeta and Jbeta gene segments in Ebeta-/- thymuses at about one-third to one-thirtieth the level found in thymuses with an unaltered TCRbeta locus. These sites are also subject to in vitro cleavage by the V(D)J recombinase in both Ebeta-/- and Ebeta+/+ thymocyte nuclei. However, the corresponding Dbeta-to-Jbeta coding joints are further reduced (by 100- to 300-fold) in Ebeta-/- thymuses. Formation of extrachromosomal Dbeta-to-Jbeta signal joints appears to be intermediately affected and nonstandard Dbeta-to-Dbeta joining occurs at the Ebeta-deleted alleles. These data indicate that, unexpectedly, loss of accessibility alone cannot explain the loss of TCRbeta recombination in the absence of the Ebeta element and suggest an additional function for Ebeta in the process of DNA repair at specific TCRbeta sites during the late phase of the recombination reaction.
    Genes & Development 09/1998; 12(15):2305-17. DOI:10.1101/gad.12.15.2305 · 12.64 Impact Factor
  • Advances in Immunology 02/1998; 69:309-52. DOI:10.1016/S0065-2776(08)60610-0 · 5.53 Impact Factor

Publication Stats

357 Citations
69.53 Total Impact Points

Institutions

  • 1998–2003
    • French Institute of Health and Medical Research
      • Immunology Research Center Marseille Luminy
      Lutetia Parisorum, Île-de-France, France
    • Centre d'Immunologie de Marseille-Luminy
      Marsiglia, Provence-Alpes-Côte d'Azur, France