Les glycolipides polysoprénoïdes et le glycosylation des protéines au niveau du reticulum endoplasmique rugueux du foie

Thèse de doctorat en sciences (chimie physiologique) (BCHM) -- UCL, 197

Identification and characterization of the tumor-specific P1A gene product

In murine mastocytoma P815, gene P1A directs the expression of antigens P815A and B which are the target of a T cell-mediated rejection response in syngeneic animals. This gene is expressed at a high level in various tumors, but is silent in normal tissues except testis and placenta; its activation is thus possibly related to malignant transformation. An anti-synthetic peptide rabbit antiserum reacted by immunoblotting with a cellular protein migrating near 40 kDa on SDS-PAGE. The immunoreactive protein was detected only in lysates from cells which express antigen P815A: P1.HTR mastocytoma cells and, after transfection with cosmids carrying the P1A gene, the antigen-loss variant P0.HTR cells and DAP-3 H-2Ld fibroblasts. The identity of this protein as the P1A gene product was confirmed by cell-free transcription-translation of the P1A cDNA, the product of which also migrated near 40 kDa in SDS-PAGE and was captured by protein A-Sepharose in the presence of the antiserum. Subcellular fractionation by differential and isopycnic centrifugation indicated that the P1A protein is associated with cytoplasmic membranes demonstrating a broad distribution with respect to size and density. Immunofluorescence microscopy also revealed a cytoplasmic signal, particularly intense in small vesicles, which coincides with that produced by an anti-mouse type I collagen guinea pig antiserum except near the cell periphery where the P1A signal is weaker. We conclude that the P1A protein is bound to membranes of the secretory pathway, at a concentration which goes increasing from the endoplasmic reticulum to secretion vesicles.(ABSTRACT TRUNCATED AT 250 WORDS

CD45RA on human CD8 T cells is sensitive to the time elapsed since the last antigenic stimulation.

The expression of CD45RA on CCR7- human CD8+ memory T cells is widely considered to be a marker of terminal differentiation. We studied the time course of CD45RA and CCR7 expression on human antitumoral cytotoxic T lymphocyte (CTL) clones and blood CD8+ T cells after antigenic stimulation. Our results indicate that CD45RA+ CCR7- CD8+ T cells are resting memory cells that, upon antigenic stimulation and during the next 10 days, proliferate, lose CD45RA, and transiently acquire CCR7. In the absence of further antigenic stimulation, they progressively re-express CD45RA and become CD45RA+ CCR7-. We conclude that the expression of CD45RA on these cells is indicative of the time elapsed since the last antigenic stimulation rather than the incapacity to proliferate or particularly high lytic potential. This concept leads to a reinterpretation of the significance of the presence of CD45RA+ CD8+ memory cells in patients affected by viral infections or by cancer

The tumor protein MAGE-1 is located in the cytosol of human melanoma cells

The MAGE-1 protein has been localized to the cytosol of human melanoma culture cells (MZ2-MEL.43) by subcellular fractionation. Its distribution between nuclear, mitochondrial, microsomal and cytosolic fractions was established by SDS-PAGE and immunoblotting with a rabbit antiserum and compared to that of markers for the main cell compartments. The immunoreactive material was recovered in the cytosolic fraction as a polypeptide migrating at 42 kDa, which has been further identified as the MAGE-1 protein because it comigrates with the product of cell-free transcription-translation of the MAGE-1 cDNA. Location to the cytosol was confirmed by immunofluorescence microscopy

A new tumor-specific antigen encoded by MAGE-C2 and presented to cytolytic T lymphocytes by HLA-B44.

A panel of cytolytic T lymphocyte (CTL) clones was isolated from metastases and blood samples of a melanoma patient vaccinated with MAGE-3.A1-pulsed autologous dendritic cells. We report here the identification of a new antigen encoded by the MAGE-C2 cancer-germline gene. This antigen is recognized by some of these CTL on HLA-B*4403. The sequence of the peptide is SESIKKKVL. It is processed in various melanoma cell lines expressing MAGE-C2 and HLA-B*4403. Because of the expression pattern of gene MAGE-C2, this new antigen is strictly tumor-specific and could therefore be used for peptide-based antitumoral vaccination