No abstract

SummaryMurine monoclonal antibodies (mAbs) M38 and L31 define two epitopes of a surface protein of activated lymphocytes and monocytes. It has been shown that M38 also defines a crossreactive epitope of human immunodeficiency virus type 1 (HIV1) gp120 (Beretta et al ., 1987. Eur. J. Immunol. 17:1793 . The mAb inhibits syncytia formation driven by HIV-1-infected cells. The surface protein was demonstrated to be a class I MHC ot chain, by sequence analysis of the corresponding cDNA and by immunological means. The epitopes defined by mAbs M38 and L31 are monomorphic and hidden (i.e., inaccessible to antibodies) on native HLA molecules expressed by resting cells, but can be evidenced on denatured proteins by Western blot analysis. The two epitopes become accessible after activation processes havebeen implemented, likely reflecting a conformational alteration of at chains (such as that described by Schnabl et al. 1990. J Exp. Med. 171:1431) . Consistent with molecular data are the results offunctional analysis, which indicate that the molecule recognized by M38 and L31 is a gate for pleiotropic negative signals, since the two mAbs were shown to inhibit monocyte antigen presentation and lymphocyte mitogenic proliferation, respectively.

Flow cytometry with the specific monoclonal antibody (MoAb) L31 was used to analyse the expression of HLA class I heavy chains not bound with beta 2-microglobulin (beta 2m) by neuroblastoma (NB) cell lines IMR-32 and LA-N-1. The cells, which express barely detectable amounts of beta 2m-free (L31-positive molecules) and beta 2m-complexed HLA class I antigens (W6.32- and BBM.1-reactive molecules), expressed MHC class I molecules not bound to light chains upon differentiation with either retinoic acid or serum starvation. The expression was not accompanied by an increase of surface heterodimers. Conversely, recombinant interferon-gamma (rIFN-gamma) treatment led IMR-32 and LA-N-1 cells to almost exclusively express beta 2m-complexes HLA class I heavy chains. Surface beta 2m-free MHC class I molecules displayed a molecular mass of approximately 45 kDa and did not bind exogenously added beta 2m. No changes in the synthesis of either HLA class I and beta 2m mRNAs or of L31 proteins were observed in differentiated NB cells, thus suggesting that the surface exposure of unusual HLA class I antigens is regulated post-translationally. These findings indicate that, in addition to activated lymphocytes, the surface expression of beta 2m-free class I heavy chains is a feature of other cell types, such as NB cells.

The aim of this study was to investigate the presence and the fine specificity of anti-CD4 autoantibodies in seronegative subjects sexually exposed to HIV-1. Anti-CD4 autoantibodies were previously detected in a fraction of HIV-1-seropositive individuals. Whole sera, purified IgG fractions, and supernatants of EBV-transformed lymphoblastoid cell lines were analyzed by means of ELISA, Western blot, and by competition assays using monoclonal antibodies with known fine specificities. Anti-CD4 antibodies were found in 6 of 18 individuals exposed to HIV-1 infection and who have been persistently seronegative. These antibodies inhibited HIV-1-driven syncytium formation, did not interfere with the CD4-gp120 interaction, and competed for CD4 binding with two of three anti-CD4 monoclonals with known fine specificities. Moreover, autoantibodies with the same fine specificities were found in the supernatants of oligoclonal EBV-transformed B cell lines derived from these individuals. At variance, in the HIV-1-positive patients included in our study, the anti-CD4 antibody response was directed to a broader panel of epitopes, including those involved in CD4-gp120 interactions. In conclusion, anti-CD4 antibodies specific for defined epitopes of the CD4 molecule are generated in the course of an early immune response to HIV-1 antigens in the absence of other signs of infection, as they can be detected by conventional methods. These autoantibodies may play a protective role either alone or in association with other cellular and humoral factors.

Although selected chemokines act as natural inhibitors of human immunodeficiency virus (HIV) infection, their inherent proinflammatory activity may limit a therapeutic use. To elucidate whether the antiviral and signaling functions of RANTES can be dissociated, several recombinant analogues mutated at the N terminus were generated and functionally compared with the wild‐type (WT) molecule, as well as with three previously described mutants. Substitution of selected residues within the N‐terminal region caused a marked loss of antiviral potency. By contrast, two unique analogues (C1.C5‐RANTES and L‐RANTES) exhibited an increased antiviral activity against different CXCR4‐negative HIV‐1 isolates grown in primary mononuclear cells or in macrophages. This enhanced HIV‐blocking activity was associated with an increased binding affinity for CCR5. Both C1.C5‐RANTES and L‐RANTES showed a dramatically reduced ability to trigger intracellular calcium mobilization via CCR3 or CCR5, while potently antagonizing the action of the WT chemokine. By contrast, two previously described analogues (RANTES3–68 and AOP‐RANTES) maintained a WT ability to trigger CCR5‐mediated signaling, while a third one (RANTES9–68) showed a dramatic loss of antiviral activity. These data demonstrate that the antiviral and signaling functions of RANTES can be uncoupled, opening new perspectives for the development of chemokine‐based therapeutic approaches for HIV infection.