Flavia Bottarel

The T cell activation molecule H4 and the CD28-like molecule ICOS are identical.

The recently cloned CD28‐like molecule ICOS displays striking similarities with H4, characterized some years ago in the mouse and recently in humans. Both molecules are selectively expressed by activated and germinal center T cells, display similar structure, and display co‐stimulatory activities. H4 displays lateral association with the CD3/TCR and is expressed by mature thymocytes. In the mouse, H4 is also expressed at high levels by thymic NKT cells that are resistant to negative selection. The aim of this work was to evaluate whether H4 and ICOS are the same molecule using the C398.4A (binding human and mouse H4) and F44 (binding human ICOS) monoclonal antibody (mAb) in parallel experiments on human T cells. ICOS and H4 displayed the same expression pattern in a panel of T cell lines and the same expression kinetics in phytohemagglutinin‐activated T cells. C398.4A completely blocked cell staining by F44, whereas F44 partially blocked C398.4A. H4 and ICOS immunoprecipitates displayed identical SDS‐PAGE patterns and H4 immunoprecipitation completely removed ICOS from cell lysates. Finally, the C398.4A mAb specifically stained cells transfected with the human or mouse ICOS. These data prove that H4 and ICOS are the same molecule and that F44 and C398.4A bind partially different epitopes.

Human CD38 interferes with HIV-1 fusion through a sequence homologous to the V3 loop of the viral envelope glycoprotein gp120.

CD38 is a progression marker in HIV‐1 infection, it displays lateral association with CD4, and down‐modulates gp120/CD4 binding. The aim of this study was to elucidate the mechanism behind the interplay between CD4, CD38, and HIV‐1. We used mouse cell transfectants expressing human CD4 and either CD38 or other CD4‐associated molecules to show that CD38 specifically inhibits gp120/CD4 binding. Human cell transfectants expressing truncated forms of CD38 and bioinformatic analysis were used to map the anti‐HIV activity and show that it is concentrated in the membrane‐proximal region. This region displayed significant sequence‐similarity with the V3 loop of the HIV‐1 gp120 glycoprotein. In line with this similarity, synthetic soluble peptides derived from this region reproduced the anti‐HIV effects of full‐length CD38 and inhibited HIV‐1 and HIV‐2 primary isolates from different subtypes and with different coreceptor use. A multiple‐branched peptide construct presenting part of the sequence of the V3‐like region potently and selectively inhibited HIV‐1 replication in the nanomolar range. Conversely, a deletion in the V3‐like region abrogated the anti‐HIV‐1 activity of CD38 and its lateral association with CD4. These findings may provide new insights into the early events of HIV‐1 fusion and strategies to intervene.

Expression of the Novel T Cell Activation Molecule hpH4 in HIV-Infected Patients: Correlation with Disease Status

We have described hpH4, a surface glycoprotein selectively expressed by activated T cells and mature thymocytes and displaying weak lateral association with CD4. The hpH4 expression pattern and biochemical features, together with analysis of its tryptic digest by peptide mass searching using MALDI-MS, suggested that it is a novel molecule. The aim of this work was to evaluate the peripheral blood T cell expression of hpH4 in HIV-infected patients and the interplay between HIV gp120 and hpH4, since both molecules interact with CD4. hpH4 expression during HIV-1 infection was evaluated by assessing 55 patients at various disease stages and following up 3 patients with primary infection and 3 patients with AIDS. hpH4 expression displayed a peak in the early phase of primary infection, dropped to control levels in the asymptomatic phase, and was newly expressed, at low levels, as AIDS developed. The expression kinetics were different than those shown by HLA-DR, CD25, and CD38. The most striking findings were the transient hpH4 expression peak displayed in the earliest stage, which was unique for hpH4. Incubation of T cells from normal donors with HIV gp120 induced transient hpH4 expression in resting CD4+ T cells and potentiated the hpH4 lateral association with CD4 in activated T cells. Moreover, hpH4 triggering inhibited gp120-induced death of CD4+ cells. Therefore, H4 expression may be a response to avoid apoptosis induced by HIV products.

Effects of the human CD38 glycoprotein on the early stages of the HIV-1 replication cycle

CD38 displays lateral association with the HIV‐1 receptor CD4. This association is potentiated by the HIV‐1 envelope glycoprotein gp120. The aim of this work was to evaluate the CD38 role in T cell susceptibility to HIV‐1 infection. Using laboratory X4 HIV‐1 strains and X4 and X4/R5 primary isolates, we found that CD38 expression was negatively correlated to cell susceptibility to infection, evaluated as percentage of infected cells, release of HIV p24 in the supernatants, and cytopathogenicity. This correlation was at first suggested by results obtained in a panel of human CD4+ T cell lines expressing different CD38 levels (MT‐4, MT‐2, C8166, CEMx174, Supt‐1, and H9) and then demonstrated using CD38 transfectants of MT‐4 cells (the line with the lowest CD38 expression). To address whether CD38 affected viral binding, we used mouse T cells that are non‐permissive for productive infection. Gene transfection in mouse SR.D10.CD4~.F1 T cells produced four lines expressing human CD4 and/or CD38. Ability of CD4+CD38+ cells to bind HIV‐1 or purified recombinant gp120 was significantly lower than that of CD4+CD38~ cells. These data suggest that CD38 expression inhibits lymphocyte susceptibility to HIV infection, probably by inhibiting gp120/CD4‐dependent viral binding to target cells.—Savarino, A., Bottarel, F., Calosso, L., Feito, M. J., Bensi, T., Bragardo, M., Rojo, J. M., Pugliese, A., Abbate, I., Capobianchi, M. R., Dianzani, F., Malavasi, F., and Dianzani, U. Effects of the human CD38 glycoprotein on the early stages of the HIV‐1 replication cycle. FASEB J. 13, 2265–2276 (1999)

The cell death-inducing ability of glycoprotein 120 from different HIV strains correlates with their ability to induce CD4 lateral association with CD95 on CD4+ T cells.

CD4 cross-linking by HIV gp120 triggers CD4+ T cell death. Several authors have suggested that this effect is mediated by CD95, but this possibility is debated by other authors. In a previous work, we found by co-capping that gp120(451) and gp120MN, but not gp120(IIIB), induce lateral association of CD4 with CD95 on the T cell surface. In this work, we used fluorescence resonance energy transfer to confirm that CD4/CD95 lateral association is induced by gp120(451), but not gp120(IIIB). Moreover, we found that gp120 ability to induce the CD4/CD95 association correlates with ability to induce cell death, since gp120(451) and gp120MN induced higher levels of cell death than did gp120(IIIB) in PHA-derived CD4+ T cell lines. CD95 involvement in gp120-induced cell death was confirmed by showing that gp120(451) and gp120MN did not induce death in CD4+ T cells derived from patients with autoimmune/lymphoproliferative disease (ALD) and decreased CD95 function. Cell death induced by gp120MN was inhibited by a recombinant CD95/IgG.Fc molecule blocking the CD95/CD95L interaction. However, inhibition was late and only partial. These data suggest that the gp120-induced CD4/CD95 association exerts a dual effect: an early effect that is independent of CD95L and may be due to direct triggering of CD95 by gp120, and a late effect that may be due to sensitization of CD95 to triggering by CD95L. In line with the former effect, cell treatment with gp120MN activated caspase 3 in the presence of Fas/IgG.Fc, which shows that cell death induced by gp120MN independently of CD95L uses the same pathway as CD95.