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P Benaroch

Publications and source records attributed to P Benaroch.

16 recordsLinked to original sources
PubMed1994

Mapping functional regions in the lumenal domain of the class II-associated invariant chain.

The MHC class II-associated invariant chain interacts in trimeric form with class II molecules, inhibits peptide binding, and mediates targeting of class II molecules to endosomal compartments. To dissect the different functions of the invariant (Ii) chain, a set of cDNAs, encoding truncated forms of the Ii chain, was constructed. mRNAs, transcribed from these cDNAs were translated in vitro, together with mRNAs encoding class II HLA DR1 alpha and beta subunits. An Ii chain truncation that contains the 104 NH2-terminal amino acids was able to associate with class II molecules. This construct contains the region from which class II-associated Ii chain peptides (CLIP, amino acids 81-104) are derived. The absence of a further eight residues at the COOH terminus results in a construct of 96 amino acids that is unable to associate with class II molecules. Association of the truncated Ii chains with class II molecules showed a strict correlation with inhibition of peptide binding. Removal of the NH2-terminal cytoplasmic tail and transmembrane region of Ii chain and its replacement with a cleavable signal sequence led to aberrant folding and impaired association with class II molecules. The region between amino acids 163 and 183 was found to be essential for visualization of Ii chain homotrimers by covalent cross-linking.

Amino Acid Sequence↗
PubMed1994

Assembly of HLA DR1 molecules translated in vitro: binding of peptide in the endoplasmic reticulum precludes association with invariant chain.

MHC class II molecules usually bind peptides in the endocytic pathway, but can also present endogenous peptides from newly synthesized proteins in a chloroquine-insensitive manner, suggesting that peptide binding might occur in the endoplasmic reticulum (ER). We used in vitro translation of HLA-DR1 class II molecules in the presence of microsomes to study peptide binding in the ER. Formation of functional class II molecules in vitro depends on formation of disulfide bridges in alpha and beta chains. The class II alpha beta heterodimers made by in vitro translation resemble class II molecules synthesized in cells in (i) their reactivity with conformation-specific antibodies, (ii) their assembly with Ii chain homotrimers, (iii) the generation of SDS-stable dimers upon peptide binding and (iv) their specificity of peptide binding. The assembly of class II molecules occurs via an alpha beta intermediate and can occur post-translationally, but only in intact microsomes. Class II alpha beta heterodimers are able to bind peptides in ER-derived microsomes, a process that precludes subsequent association of class II molecules with Ii chain. This mechanism might explain presentation of endogenous peptides by class II molecules.

Antibodies, Monoclonal↗
PubMed1994

T cell-induced Ig allotypic suppression in mice. Basis for emergence or tolerization, during the perinatal period, of natural T cells specific to the IgG2ab allotype.

We have previously described an anti-IgG2ab T cell activity in normal Igha/a mice. Their congenic partner at the Igh locus (Ighb/b) and Igha/b hybrids bred from them, do not display this T cell activity but express IgG2ab. As these mice are supposed to possess the same genetic elements related to this potential T cell repertoire, only somatic selection mechanisms could be responsible for their different behavior. In this study, we investigated the basis for the emergence (in Igha/a mice) or tolerization (in Ighb/b-congenic mice and in Igha/b hybrids) of these natural anti-IgG2ab T cells. Stringent perinatal B cell deprivation in Ighb/b and Igha/b mice resulted in the emergence of anti-IgG2ab T cells, as these individuals could be subjected to autoimmune, T cell-mediated IgG2ab suppression. Furthermore, the acquisition of anti-IgG2ab T cell activity was drastically reduced in Igha/a mice, perinatally exposed to IgG2ab; thus, the presence of this allotype leads to tolerization of these specific T cells.

Animals↗
PubMed1993

Role of CD4+ and CD8+ cell subsets during amplification of natural T cell activity against IgG2ab in Igha mice and during induction of IgG2ab allotype suppression in Igha/b mice.

Transfer into F1 Igha/b mice of splenocytes from Igha mice sensitized once against B cells from an Ighb congenic strain induces total, chronic, and IgG2ab (IgG2a of the Ighb haplotype)-specific allotype suppression in these recipients. We previously demonstrated that both the CD4+ and CD8+ T subsets were necessary for inducing suppression, but that CD8+, cells by themselves were sufficient for maintaining suppression. We have studied the suppression induction capacity of different mixtures of CD4+ and CD8+ subsets obtained by in vitro cytotoxic treatment of T splenocytes from normal or sensitized Igha mice, and we have established that suppression induction requires the cooperation between CD4+ and CD8+ populations, both of which have to be IgG2ab specific. In addition, Igha mice were sensitized in the absence of CD4+ or CD8+ cells by in vivo cytotoxic treatment performed before and after the sensitization in order to obtain an IgG2ab-specific CD4+ population that has arisen in the absence of CD8+ cells, and vice versa. We found that only IgG2ab-specific CD4+ cells from anti-CD8-treated mice (T'sens CD4+) had the ability to induce suppression in F1 Igha/b hosts. Nevertheless, the real effector cells in this suppression model display the CD8+ phenotype, as in vivo cytotoxic anti-CD8 treatment of Igha/b recipients of T'sens CD4+ abrogates the suppression induction capacity. Taken together, these results show that T'sens CD4+ have an important capacity to recruit CD8+ anti-IgG2ab effector cells from precursors that have been transferred with them into Igha/b hosts. These precursors are actually derived from the T'sens CD4+ cell preparation, because we have recently demonstrated that suppression is maintained by donor T cells throughout the recipient's life. CD4+ cells can have their anti-IgG2ab activity amplified only by means of target cells (i.e., B cells from Ighb congenic mice), whereas, in the absence of CD4+ cells, and despite the presence of target cells, CD8+ cells seem unable to acquire this amplified activity.

Animals↗
PubMed1993

Identification of T cell peptides from C gamma 2ab involved in IgG2ab T cell-induced allotypic suppression. Relationship with the MHC haplotype of the suppressor T cell-donor Igha mice.

Using the algorithm of the peptidic self-model we selected, from the constant part of the IgG2ab polypeptidic heavy chain, four peptides susceptible to being recognized by the T cells of Igha mice that prevent IgG2ab expression. Among these four peptides, two were capable of enhancing in vivo this T cell activity of Igha mice almost as well as the whole IgG2ab allotype. One of these peptides, C gamma 2ab-248-263, located in the CH3 domain of IgG2ab, was effective in H-2d mice, while the other one, C gamma 2ab-103-118, located in the hinge region of IgG2ab, was efficient in H-2b mice. This demonstrates for the first time that this suppression involves the TCR of the effector cells and peptides derived from the allotype presented in the context of MHC molecules of the target cells, the presentation specificity being dependent on the MHC haplotype. The availability of these peptides will allow us to further understand the role of class I and/or class II MHC molecules in this suppression.

Amino Acid Sequence↗
PubMed1992

T cell-induced allotypic suppression. Origin of the CD8+ T cells maintaining IgG2ab suppression.

One of the problems raised by the T cell-induced allotypic suppression is the origin (donor or host) of the T cells responsible for the chronicity of the suppression. To address this point, we used T cells from Igha/a Thy-1.2 mice whose natural T cell activity against IgG2ab was enhanced in vivo. These T cells were injected into newborn Ighb/b Thy-1.1 mice where they induced complete suppression of IgG2ab expression in around 70% of these recipients. During a study that lasted more than 1 yr, we found that about 3% of the recipient splenocytes were T cells of the donor type. By means of suppression-transfer experiments, using either Thy-1.2+ or Thy-1.1+ cell-depleted splenocytes from mice suppressed in this manner we were able to unambiguously show that Thy-1.2+ cell-depleted splenocytes were incapable of transferring the suppression, whereas Thy-1.1+ cell-depleted splenocytes could. We thus demonstrated that suppression was maintained throughout the recipient's life by donor Thy-1.2+ T cells.

Animals↗
PubMed1992

Tetrameric cell-surface MHC class I molecules.

Purified major histocompatibility complex (MHC) class I molecules have been studied at high resolution by X-ray crystallography; the structure is a complex of a single heavy chain, a beta 2-microglobulin light chain and a tightly bound peptide moiety. We show here that complete MHC class I molecules are post-translationally assembled into tetramers (made up of four heavy chains and four beta 2-microglobulin units) and that this tetrameric species is expressed on the cell surface. The multivalent tetrameric structure of class I molecules can be reconciled with models of T-cell activation that invoke antigen-receptor crosslinking, as opposed to models that depend on an allosteric change.

Animals↗
PubMed1992

Plasmodium chabaudi and Trypanosoma cruzi infections can reverse IgG2ab chronic suppression in mice.

The effects of infections with Plasmodium chabaudi or Trypanosoma cruzi on chronic CD8+ T cell dependent IgG2ab suppression were analyzed in homozygous Ighb/b adult mice. These parasites are known to induce a CD4+ T cell dependent polyclonal activation characterized in particular by a considerable increase in IgG2a expression. We report here that infection with either parasite reversed the IgG2ab suppression in 18 out of 32 mice. However, in mice treated with anti-CD4 mAb in parallel to the parasite infection, the polyclonal activation was largely reduced and the suppression of IgG2ab expression was always maintained. The parasite induced escape from suppression could result from increased helper T cell activation stimulating some of the Ig production. A weakening of the CD8+ T cell activity which is specific of IgG2ab could also contribute to the IgG2ab production. Both effects would shift the previous equilibrium which was in favour of suppression to a new balance allowing expression of the IgG2ab allotype.

Animals↗
PubMed1989

T cell-induced Ig allotypic suppression in mice. II. Both CD4+ CD8- and CD4- CD8+ T cell subsets from sensitized Igha mice are required to induce suppression of Igh-1b allotype expression.

We established the phenotype of T splenocytes (Ts) from Igha/a BALB/c mice sensitized against B splenocytes from the Ighb/b CB20 congenic mice that induce Igh-1b (IgG2a of the Ighb haplotype) suppression. This was achieved by studying the action of anti-T cell subset mAb on the capacity of Ts to induce this chronic allotypic suppression in Igha/b (BALB/c x CB20)F1 mice. The Ts were treated with cytotoxic anti-mouse CD4 or anti-mouse CD8 rat mAb in vitro before their injection into the Igha/b newborns or in vivo after their injection into the Igha/b newborns. Exposure to either anti-CD8 or anti-CD4 mAb in vitro or in vivo leads to loss of the capacity of Ts to induce Igh-1b allotypic suppression. Mixing CD4+-cell-depleted Ts and CD8+-cell-depleted Ts preparations restored the capacity of the cells to induce Igh-1b suppression. Thus, both CD4+ CD8- Ts and CD4- CD8+ Ts are required for the induction of this allotypic suppression. Bone marrow cells and B splenocytes from Igh-1b-suppressed adult Igha/b mice were shown to be able to durably express Igh-1b when transferred into irradiated Igha/a BALB/c hosts whereas whole spleen cells from such donors failed to do it. Abrogation of Igh-1b suppression by in vivo anti-CD8 mAb treatment was achieved in adult Igha/b heterozygotes but with a lower efficiency than in adult Ighb/b homozygotes, all being chronically Igh-1b suppressed. The CD4- CD8+ cell population essential for maintaining this suppression is resistant to in vivo 600 rad irradiation and seems to be slightly inhibited by in vivo administration of free Igh-1b.

Animals↗
PubMed1988

Cellular induction of chronic allotype suppression of IgG2a in Ighb/b homozygous mice and its abrogation by in vivo treatment with anti-CD8 monoclonal antibody.

We report here the successful induction of allotype suppression in homozygous Ighb/b mice (CB20 or C57BL/6) by neonatal injection of T splenocytes from Igha congenic sensitized mice (BALB/c or BC8, respectively). The sensitization of the T cell donors was achieved by two intravenous injections of B splenocytes from Ighb congenic mice. Treated homozygous Ighb/b mice developed, as of 16-24 wk of age, a chronic suppression of Igh-1b expression (IgG2a of Ighb haplotype). The other productions tested (IgM, IgD, and IgA) of Ighb haplotype were unaffected. In vivo treatment with cytotoxic anti-CD4 or anti-CD8 mAb of mice subjected to chronic Igh-1b suppression clearly showed that CD8+ lymphocytes (suppressor or cytotoxic cell) were essential for the maintenance of the suppression. The suppression was indeed abrogated after a 1-wk treatment with anti-CD8 mAb containing culture supernatant, whereas, the anti-CD4-treated mice continued to be subjected to suppression. This anti-CD8 in vivo treatment was shown to have no effect on thymus but to severely reduce the percentages of CD8+ cells in spleen and in peripheral blood without affecting the percentages of CD4+ cells, leading to a large and rapid Igh-1b expression (up to 0.5 mg per ml of serum, the day after the end of the treatment). This suppression abrogation, and thus the Igh-1b expression, was either transient or permanent. When it was transient, a second 1-wk treatment with anti-CD8 mAb containing culture supernatant induced once again a rapid and significant production of Igh-1b (up to 0.3 mg of Igh-1b per ml of serum).

Animals↗
PubMed1988

Enhancement in Igha mouse strains of the "natural" suppressive activity of normal T splenocytes against the expression of Igh-1b allotype. I. Molecular aspects of the chronic suppression obtained.

Several approaches have been used in our attempts to increase the "natural" ability of normal T splenocytes (Tn) from BALB/c or BC8 mice (both Igha) to induce, in F1 hybrids, a suppression of Igh-1b expression (IgG2a of b haplotype). These heterozygous F1 were produced by mating these Igha mice and their Ighb-congenic partners (CB20 and C57BL/6, respectively). The most powerful approaches were to sensitize the Igha mice by either autologous splenocytes coated with Igh-1b or B splenocytes from Ighb-congenic mice. In F1 having paternally inherited the b haplotype the sensitized T splenocytes (Ts) prepared from such mice are able to induce, like Tn, when injected at birth, a chronic suppression of Igh-1b expression. However, the suppression was established with a much higher efficiency: already at 6 weeks of age in 100% of the F1 treated with 1 x 10(7) Ts, vs. a final rate of 70% progressively reached only at 42 weeks of age in the F1 treated with 4 x 10(7) Tn. In F1 having maternally inherited Ighb the differences were even more pronounced than with 4 x 10(7) Tn, i.e. the suppression induction was almost totally ineffective, whereas with 2 x 10(7)-4 x 10(7) Ts, a rate of 100% treated F1 subjected to suppression was reached at 19 weeks of age. As the productions of IgM, IgD and IgA of the b haplotype were not affected by the suppression, the Ts are believed to act on the Igh-1b+ cells. Attempts were also made to induce allotypic suppression of other b allotypes by the use, as sensitizing cells, of myeloma cells carrying Igh-6b (IgM of b haplotype). We failed in revealing any sign of a T cell reactivity against Igh-6b similar to the reactivity against Igh-1b. The use of Igh-6b+ myeloma cells grown in an Ighb or in an Igha background allowed us to assume that the cells responsible for the sensitization are, in the Ighb B lymphocyte population, either the Igh-1b+ lymphocytes or the lymphocytes having passively adsorbed this allotype, or both.

Animals↗
PubMed1987

Normal T splenocytes are able to induce immunoglobulin allotypic suppression in F1 hybrid mice.

A chronic suppression of Igh-1b and Igh-3b (IgG2a and IgG2b of b haplotype) allotype expression has been induced by injecting T splenocytes from normal BALB/c or BC8 mice into newborn F1 hybrids of appropriate Igh congenic strains: BALB/c into (BALB/c Igha X CB20 Ighb)F1 and BC8 into (BC8 Igha X C57BL/6 Ighb)F1 or (C57BL/6 X BC8)F1. This suppression does not affect IgM (IgH-6b) or IgA (Igh-2b) expression. When the Ighb haplotype is paternally transmitted, the proportion of T splenocyte recipients showing allotypic suppression increases with time reaching 70% 40 weeks after birth. We also succeeded in inducing this pattern of suppression in 2 out of 13 cases when the Ighb was inherited from the mother. These normal T splenocytes are therefore clearly allotype specific. As Igh-6b production is not affected by the suppression, these T splenocytes are believed to influence B cells more or less committed to Igh-1b or Igh-3b production rather than more precocious Igh-6b (IgM of b haplotype) carrying precursors in the classical IgM-IgG filiation pathway.

Animals↗
PubMed1984

Induction of immunoglobulin allotypic suppression in the rabbit: a cellular approach.

Adult rabbits were sensitized against rabbit immunoglobulin allotypic specificities of the a and b series by means of two injections of their own lymphocytes coated with the appropriate IgG. The splenic lymphocytes from these sensitized rabbits were injected, 24 hr after birth, into newborns that had inherited from their father the allotypic specificities susceptible to recognition by the sensitized lymphocytes. A significant percentage (40%) of the young rabbits subjected to this treatment developed a total suppression of expression of these specificities with the well-known compensatory effect due to an increased expression of the corresponding allele. We have observed that the T Julius subset of the sensitized splenic lymphocytes was responsible for the induction of this suppression that was established without discernible intervention of anti-allotype antibodies. In one instance we observed that the expression of the allotypic specificities transmitted by the father, although established at 5 weeks of age, gradually declined and totally disappeared at 13 weeks of age.

Animals↗