Biomedical subjects

N Auphan

Publications and source records attributed to N Auphan.

22 records · Page 2Linked to original sources

PubMed1991

Biochemical and functional association between CD8 and H-2 at the surface of a T cell clone.

In an attempt to define structures interacting with CD8 molecules during activation of CD8+ cells, immunoprecipitates of CD8 and Tcr-CD3 molecules from lysates of a surface-labeled CTL clone were analyzed. No proteins other than the known Tcr alpha/beta and associated CD3 components were detected in either anti-Tcr or anti-CD3 immunoprecipitates, whether or not the CTL clone had been activated. However, anti-CD8 antibodies co-precipitated class I MHC heavy chain and associated beta 2-microglobulin in all conditions. The latter co-precipitation was shown to result from "cis-type" interactions between CD8 and class I MHC proteins on the same cell and to involve a degree of selectivity, as class I MHC molecules were absent from immunoprecipitates of highly expressed cell surface molecules such as LFA-1. A further analysis of cell surface molecular distribution during antigen-dependent CTL-target cell interaction by double fluorescence-microscopy in non-activating conditions indicated that an increased density of CTL class I molecules was found in the CTL-target cell contact zone of most conjugates with redistributed CD8 molecules. A possible role for "cis-type" class I MHC-CD8 interactions in the dynamics of CTL-target cell contacts is proposed.

Animals↗

PubMed1991

Non-deletional mechanisms of peripheral and central tolerance: studies with transgenic mice with tissue-specific expression of a foreign MHC class I antigen.

The studies described here reveal a surprising variety of non-deletional manifestations of tolerance (anergy and unresponsiveness) in both the thymus and peripheral lymphoid organs. This can be observed in anti-Kb TCR transgenic mice crossed with normal Kb positive mice, and in TCR mice crossed with transgenic mice expressing Kb in restricted tissues, such as liver, epithelial cells, cells of neuroectodermal origin etc. Thymic induction of unresponsiveness: Transgenic mice were prepared with the a, beta TCR genes from a CD8-dependent and Kb-specific CTL clone. In homozygous H-2b mice clonotype+ cells were found in the thymus but none or few in the periphery, suggesting that deletion occurs in the thymus although lack of migration to the periphery has not been ruled out. In H-2 heterozygous mice (TCR.H-2kxb) deletion was incomplete in the thymus and clonotype+ cells accumulated substantially in the periphery. Most of them had downregulated their CD8 molecules. The clonotype+ cells in both the thymus and periphery were unresponsive to the Kb antigen in vitro, suggesting the induction of anergy in the thymus. The inefficient negative selection in H-hkxb heterozygous mice is probably due to the lower Kb expression in comparison to H-2b homozygous mice. We then further reduced the amount of Kb expression in the thymus by constructing Kb transgenic mice using the 0.8 Kb fragment of the keratin IV promoter. In the thymus expression was only observed on a subset of medullary epithelial cells. When these mice were crossed with the TCR transgenic mice than no deletion was observed in the thymus. However, the clonotype+ CD8+ thymocytes could not respond to Kb in vitro, indicating that they had been rendered anergic in the thymus. These data show that unresponsiveness can be induced in the thymus, that the extent of clonal deletion can vary greatly owing to a change in antigen expression by a factor of two as in H-2 heterozygous versus homozygous mice, and that expression of Kb on a few medullary thymic epithelial cells is sufficient to induce anergy. Peripheral induction of unresponsiveness: To study the consequence of tissue-specific tolerogen expression we have generated transgenic mice expressing Kb exclusively in cells of neuroectodermal origin (GFAP.Kb mice) in the liver (alumin.Kb mice), or in certain epithelial cells outside the thymus (2.4 keratin IV.Kb mice). Consistent with the absence of Kb expression in the thymus there was no deletion of clonotype+ CD8+ cells in the thymus, and the thymocytes were fully functional.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1989

Comparison of phosphorylation and internalization of the antigen receptor/CD3 complex, CD8, and class I MHC-encoded proteins on T cells. Role of intracytoplasmic domains analyzed with hybrid CD8/class I molecules.

We analyzed the phosphorylation and the dynamics of TCR/CD3, CD8 and MHC class I molecules during the activation of a CD8+ cytotoxic T lymphocyte clone and of CD8- T helper hybridomas transfected with the gene coding for the native (J. Gabert, C. Langlet, R. Zamoyska, J.R. Parnes, A.M. Schmitt-Verhulst, and B. Malissen. 1987. Reconstitution of MHC class I specificity by transfer of the T cell receptor and Lyt-2 genes. Cell 50:545) or truncated CD8 alpha molecule. The CD3 components gamma and epsilon and the CD8 alpha subunit were phosphorylated after activation of the CTL clone with the protein kinase C activator PMA. Class I MHC molecules were phosphorylated irrespective of PMA activation. Constitutive phosphorylation of the MHC class I products was found to be intrinsic to the transmembrane/cytoplasmic portion of the molecules because it was transferred to the CD8 alpha hybrid molecules composed of extracellular CD8 and MHC class I transmembrane and intracytoplasmic domains (CD8-e/MHC-t-i). Measurements of the dynamics of these cell surface molecules by using radiolabeled mAb revealed distinct behaviors: TCR/CD3 complex ligand internalization was increased (around 50% after 40 to 60 min) after PMA activation, whereas the ligand of class I MHC molecules was internalized at constant rate irrespective of PMA activation. Ligand bound to native CD8 molecules was poorly internalized, irrespective of the activation of the T cells with PMA. The same ligand bound to the CD8-e/MHC-t-i hybrid molecule was internalized at the same rate as a class I MHC molecule ligand, indicating that the behavior of the hybrid molecule was characteristic of the transmembrane/cytoplasmic portion of MHC class I molecules.

Amino Acid Sequence↗

PubMed1989

Antibodies to HLA class I promoter-binding proteins in sera from patients with systemic lupus erythematosus.

Antibodies directed against human cell nuclear factors (HLA-F1 and HLA-F2) that bind to the interferon consensus sequence (ICS) in the promoter region of the HLA-A2 class I gene can be detected in the serum of some patients with systemic lupus erythematosus (SLE). Such antibodies, in gel-shift assays, inhibited the formation of the retarded band corresponding to the HLA-F1/F2-ICS complex irrespective whether purified factors or crude nuclear extracts were used. This inhibitory effect was due to IgG since the SLE serum activity bound to and could be eluted from a protein A-Sepharose column. This effect still persisted after absorption of SLE serum on a DNA-affinity column which removed antibodies directed against protein-free HLA-A2-ICS nucleotide sequence also present in such sera. Western blot analysis further confirmed that SLE serum recognized both HLA-F1 (65 kDa) and HLA-F2 (88 kDa) polypeptides, independently of the presence of the HLA-A2-ICS nucleotidic sequence, as well as other proteins. These results suggest that human sera from some patients with SLE may be used as a convenient source of antibodies directed against DNA-binding proteins regulating gene transcription.

DNA-Binding Proteins↗