Protection of murine systemic lupus by the Ea transgene without expression of I-E heterodimers

A high-level expression of the Ea transgene encoding the MHC class II I-E alpha-chain is very effective in the protection from systemic lupus erythematosus (SLE) in mice. However, it has not been elucidated whether this protection results from the induction or increased expression of I-E heterodimers or from the generation of I-E alpha-chain-derived peptides displaying high affinity for I-A molecules, because previous studies were conducted in lupus-prone mice expressing I-E beta-chains. To address this question, we assessed the protective effect of the Ea transgene in lupus-prone BXSB mice bearing the H2(q) haplotype (i.e., unable to express I-E heterodimers because of a deficiency in I-E beta-chains). We observed that the Ea transgene expression resulted in a marked suppression of the development of SLE in H2(q) BXSB mice despite the absence of I-E expression. The observed protection was not associated with any detectable levels of T cell depletion and regulatory T cell expansion. Significantly, transgenic I-E alpha-chains were substantially expressed on the surface of B lymphocytes and dendritic cells, but not of macrophages, without apparent formation of mixed-isotype heterodimers with I-A beta-chains. Our results demonstrate for the first time that the Ea transgene is able to prevent the development of SLE without induction of I-E heterodimer expression, indicating a critical role of I-E alpha-chains, but not I-E heterodimers, in the Ea transgene-mediated protection from SLE. Taken together, our data favor a model of autoimmunity prevention based on competition for Ag presentation between I-E alpha-chain-derived peptides and autoantigens.     

IL-12 administration reveals diabetogenic T cells in genetically resistant I-Ealpha-transgenic nonobese diabetic mice: resistance to autoimmune diabetes is associated with binding of Ealpha-derived peptides to the I-A(g7) molecule

Nonobese diabetic (NOD) and NOD-DRalpha transgenic (tg) mice, expressing Aalpha(d):Abeta(g7) and Aalpha(d):Abeta(g7) plus DRalpha:Ebeta(g7) class II molecules, respectively, both develop insulin-dependent diabetes mellitus (IDDM), whereas NOD-Ealpha tg mice expressing Aalpha(d):Abeta(g7) plus Ealpha:Ebeta(g7) are protected. We show that IL-12 administration induces rapid IDDM onset in NOD-DRalpha but fails to provoke insulitis and diabetes in NOD-Ealpha tg mice. Nevertheless, T cells from IL-12-treated NOD-Ealpha tg mice secrete IFN-gamma and transfer IDDM to NOD-SCID and NOD-Ealpha-SCID recipients, demonstrating the presence of peripheral diabetogenic Th1 cells in the protected mice. Surprisingly, regulatory cells were undetectable. Moreover, Ealpha:Ebeta(g7) could substitute for DRalpha:Ebeta(g7) in Ag presentation, arguing against mechanisms of protection involving capture of diabetogenic I-A(g7)-restricted epitopes by Ealpha:Ebeta(g7)molecules. Interestingly, the expression of naturally processed epitopes derived from DRalpha- and Ealpha-chains bound to I-A(g7) is different in the two strains of tg mice, and the difference is enhanced by IL-12 administration. I-A(g7) molecules from both NOD-DRalpha and NOD-Ealpha tg mice present the conserved DRalpha/Ealpha 52-68 sequence, at high and low levels, respectively. In addition, only IDDM-resistant NOD-Ealpha tg mice possess APCs bearing Ealpha65-77/I-A(g7) complexes, which tolerize the specific T cells. This is associated with the selective inhibition of the response to insulinoma-associated protein 2 (IA-2), an autoantigen in IDDM. Our results support protective mechanisms based on I-A(g7) blockade by peptides unique to the Ealpha-chain, such as Ealpha65-77 and/or tolerance of diabetogenic T cells cross-reactive with Ealpha-peptide/I-A(g7) complexes.     

Role of dendritic cells in the immune response induced by mouse mammary tumor virus superantigen.

After mouse mammary tumor virus (MMTV) infection, B lymphocytes present a superantigen (Sag) and receive help from the unlimited number of CD4(+) T cells expressing Sag-specific T-cell receptor Vbeta elements. The infected B cells divide and differentiate, similarly to what occurs in classical B-cell responses. The amplification of Sag-reactive T cells can be considered a primary immune response. Since B cells are usually not efficient in the activation of naive T cells, we addressed the question of whether professional antigen-presenting cells such as dendritic cells (DCs) are responsible for T-cell priming. We show here, using MMTV(SIM), a viral isolate which requires major histocompatibility complex class II I-E expression to induce a strong Sag response in vivo, that transgenic mice expressing I-E exclusively on DCs (I-EalphaDC tg) reveal a strong Sag response. This Sag response was dependent on the presence of B cells, as indicated by the absence of stimulation in I-EalphaDC tg mice lacking B cells (I-EalphaDC tg muMT(-/-)), even if these B cells lack I-E expression. Furthermore, the involvement of either residual transgene expression by B cells or transfer of I-E from DCs to B cells was excluded by the use of mixed bone marrow chimeras. Our results indicate that after priming by DCs in the context of I-E, the MMTV(SIM) Sag can be recognized on the surface of B cells in the context of I-A. The most likely physiological relevance of the lowering of the antigen threshold required for T-cell/B-cell collaboration after DC priming is to allow B cells with a low affinity for antigen to receive T-cell help in a primary immune response.     

Diverse repertoire of the MHC class II-peptide complexes is required for presentation of viral superantigens

Among other features, peptides affect MHC class II molecules, causing changes in the binding of bacterial superantigens (b-Sag). Whether peptides can alter binding of viral superantigens (v-Sag) to MHC class II was not known. Here we addressed the question of whether mutations limiting the diversity of peptides bound by the MHC class II molecules influenced the presentation of v-Sag and, subsequently, the life cycle of the mouse mammary tumor virus (MMTV). T cells reactive to v-Sag were found in mice lacking DM molecules as well as in A(b)Ep-transgenic mice in which MHC class II binding grooves were predominantly occupied by an invariant chain fragment or Ealpha(52-68) peptide, respectively. APCs from the mutant mice failed to present v-Sag, as determined by the lack of Sag-specific T cell activation, Sag-induced T cell deletion, and by the aborted MMTV infection. In contrast, mice that express I-A(b) with a variety of bound peptides presented v-Sag and were susceptible to MMTV infection. Comparison of v-Sag and b-Sag presentation by the same mutant cells suggested that presentation of v-Sag had requirements similar to that for presentation of toxic shock syndrome toxin-1. Thus, MHC class II peptide repertoire is critical for recognition of v-Sag by the T cells and affects the outcome of infection with a retrovirus.     

H2-Mbeta 1 and H2-Mbeta 2 heterodimers equally promote clip removal in I-A(q) molecules from autoimmune-prone DBA/1 mice

Antigen-presenting cells degrade endocytosed antigens, e.g. collagen type II, into peptides that are bound and presented to arthritogenic CD4(+) helper T cells by major histocompatibility complex (MHC) class II molecules. Efficient loading of many MHC class II alleles with peptides requires the assistance of H2-M (HLA-DM in humans), a heterodimeric MHC class II-like molecule that facilitates CLIP removal from MHC class II molecules and aids to shape the peptide repertoire presented by MHC class II to CD4(+) T cells. In contrast to the HLA-DM region in humans, the beta-chain locus is duplicated in mice, with the H2-Mb1 beta-chain distal to H2-Mb2 and the H2-Ma alpha-chain gene. H2-M alleles appear to be associated with the development of autoimmune diseases. Recent data showed that Mbeta1 and Mbeta2 isoforms are differentially expressed in isolated macrophages and B cells, respectively. The tissue expression and functional role of these heterodimers in promoting CLIP removal and peptide selection have not been addressed. We utilized the human T2 cell line, which lacks part of chromosome 6 encompassing the MHC class II and DM genes, to construct transgenic cell lines expressing the MHC class II heterodimer I-A(q) alone or in the presence of H2-Malphabeta1 or H2-Malphabeta2 heterodimers. Both H2-M isoforms facilitate the exchange of CLIP for cognate peptides on I-A(q) molecules from arthritis-susceptible DBA/1 mice and induce a conformational change in I-A(q) molecules. Moreover, I-A(q) cell-surface expression is not absolutely dependent on H2-M molecules. These data suggest that I-A(q) exhibits a high affinity for CLIP since virtually all I-A(q) molecules on T2 cells were found to be associated with CLIP in the absence of both H2-M isoforms.     

H2E-derived Ealpha52-68 peptide presented by H2Ab interferes with clonal deletion of autoreactive T cells in autoimmune thyroiditis

Susceptibility and resistance to experimental autoimmune thyroiditis is encoded by MHC H2A genes. We reported that traditionally resistant B10 (H2(b)) mice permit thyroiditis induction with mouse thyroglobulin (mTg) after depleting regulatory T cells (Tregs), supporting A(b) presentation to thyroiditogenic T cells. Yet, Ea(k) transgenic mice, expressing A(b) and normally absent E(b) molecules (E(+)B10 mice), are susceptible to thyroiditis induction without Treg depletion. To explore the effect of E(b) expression on mTg presentation by A(b), seven putative A(b)-binding, 15-16-mer peptides were synthesized. Five were immunogenic for both B10 and E(+)B10 mice. The effect of E(b) expression was tested by competition with an Ealpha52-68 peptide, because Ealpha52-68 occupies approximately 15% of A(b) molecules in E(+)B10 mice, binding with high affinity. Ealpha52-68 competitively reduced the proliferative response to mTg, mTg1677, and mTg2342 of lymph node cells primed to each Ag. Moreover, mTg1677 induced mild thyroiditis in Treg-depleted B10 mice, and in E(+)B10 mice without the need for Treg depletion. Ealpha52-68 competition with mTg-derived peptides may impede clonal deletion of pathogenic, mTg-specific T cells in the thymus.     

T cells that recognize peptide sequences of self MHC class II molecules exist in syngeneic mice.

T cell reactivity toward self MHC class II molecules has been recognized in syngeneic MLR in a number of studies, where the T cells are believed to recognize the combination of self/nonself peptide and self MHC molecule. We investigated the stimulation of T cell proliferation by synthetic peptides of sequences corresponding to the first polymorphic amino terminal domain of alpha- and beta-chains of self I-A molecules. Both unprimed and primed T cells responded to a number of peptides of alpha 1 and beta 1 domains of self I-Ad molecules. The response was dependent on the presentation of I-Ad peptides by syngeneic APC and was blocked by anti-class II MHC mAb. Upon further investigation it was observed that I-Ad peptides could inhibit the stimulation of Ag-specific MHC class II-restricted T cell hybridoma due to self presentation of peptides rather than to direct binding of free peptides to the TCR, further supporting their affinity/interaction with intact self MHC class II molecules. The peptide I-A beta d 62-78 showed high affinity toward intact self MHC II molecule as determined by the inhibition of Ag-specific T cell stimulation and yet was nonstimulatory for syngeneic T cells, therefore representing an MHC determinant that may have induced self tolerance. Thus we have shown that strong T cell proliferative responses can be generated in normal mice against the peptides derived from self MHC class II molecules and these cells are part of the normal T cell repertoire. Therefore complete tolerance toward potentially powerful immunodominant but cryptic determinants of self Ag may not be necessary to prevent autoimmune diseases.     

Lack of MHC-II expression in activated mouse T cells correlates with DNA methylation at the CIITA-PIII region

In contrast to activated human T cells, activated mouse T cells fail to express MHC class II molecules (MHC-II) at their cell surface. This is because mouse T cells hardly produce mRNA encoding the MHC-II molecules I-A and I-E, due to severely impaired expression levels upon T-cell activation of the mhc2ta gene, encoding the class II transactivator (CIITA). In humans, activated T cells express exclusively the CIITA promoter III (CIITA-PIII) isoform, which results in cell surface expression of all MHC-II isotypes (HLA-DR, -DP and -DQ). In this study, we demonstrate that methylation of CIITA-PIII contributes to the failure of mouse T cells to transcribe the mhc2ta and the resulting I-A/E genes, explaining the lack of I-A/E molecule expression at the cell surface following activation.     

Self-specific MHC class II-restricted CD4-CD8- T cells that escape deletion and lack regulatory activity

In the presence of the I-Ealpha protein, transgenic (Tg) mice expressing the 1H3.1 alphabeta TCR that is specific for the Ealpha52-68:I-A(b) complex display drastic intrathymic deletion. Although peripheral T cells from these mice remained unresponsive to the Ealpha52-68:I-A(b) complex, they contained a subpopulation able to specifically react to this complex in the presence of exogenous IL-2, indicating that some 1H3.1 alphabeta TCR Tg T cells have escaped clonal deletion and efficiently populated the periphery. IL-2-dependent, Ealpha52-68:I-A(b) complex-responsive T cells were CD4-CD8- and expressed the 1H3.1 alphabeta TCR. Such T cells could develop intrathymically, did not show sign of regulatory/suppressor activity, displayed a typical naive phenotype, and seemed to persist in vivo over time. CD4-CD8- TCR Tg T cells were also detected when the surface density of the deleting ligand was increased on MHC class II+ cells. In addition, the development of CD4-CD8- 1H3.1 alphabeta TCR Tg T cells could be supported by I-A(b) molecules. These observations indicate that CD4 surface expression neither specifies, nor is required for, the thymic export of mature thymocytes expressing a MHC class II-restricted alphabeta TCR. The data also show that, although the avidity of the interaction involved in intrathymic deletion is significantly lower than that involved in mature T cell activation, its range can be large enough to be influenced by the presence or absence of coreceptors. Finally, the margin created by the absence of CD4 coreceptor was substantial because it could accommodate various amounts of the deleting ligand on thymic stromal cells.     

Nonobese diabetic mice display elevated levels of class II-associated invariant chain peptide associated with I-Ag7 on the cell surface

Peptide presentation by MHC class II molecules plays a pivotal role in determining the peripheral T cell repertoire as a result of both positive and negative selection in the thymus. Homozygous I-A(g7) expression imparts susceptibility to autoimmune diabetes in the nonobese diabetic mouse, and recently, it has been proposed that this arises from ineffectual peptide binding. Following biosynthesis, class II molecules are complexed with class II-associated invariant chain peptides (CLIP), which remain associated until displaced by Ag-derived peptides. If I-A(g7) is a poor peptide binder, then this may result in continued occupation by CLIP to the point of translocation to the cell surface. To test this hypothesis we generated affinity-purified polyclonal antisera that recognized murine CLIP bound to class II molecules in an allele-independent fashion. We have found abnormally high natural levels of cell surface class II occupancy by CLIP on nonobese diabetic splenic B cells. Experiments using I-A-transfected M12.C3 cells showed that I-A(g7) alone was associated with elevated levels of CLIP, suggesting that this was determined solely by the amino acid sequence of the class II molecule. These results indicated that an intrinsic property of I-A(g7) would affect both the quantity and the repertoire of self-peptides presented during thymic selection.     

T cell reactivity to MHC class II-bound self peptides in systemic lupus erythematosus-prone MRL/lpr mice

The epitopes recognized by pathogenic T cells in systemic autoimmune disease remain poorly defined. Certain MHC class II-bound self peptides from autoimmune MRL/lpr mice are not found in eluates from class II molecules of MHC-identical C3H mice. Eleven of 16 such peptides elicited lymph node cell and spleen cell T cell proliferation in both MRL/lpr (stimulation index = 2.03-5.01) and C3H mice (stimulation index = 2.03-3.75). IL-2 and IFN-gamma production were detected, but not IL-4. In contrast to what was seen after immunization, four self peptides induced spleen cell proliferation of T cells from naive MRL/lpr, but not from C3H and C57BL/6.H2(k), mice. These peptides were derived from RNA splicing factor SRp20, histone H2A, beta(2)-microglobulin, and MHC class II I-A(k)beta. The first three peptides were isolated from I-E(k) molecules and the last peptide was bound to I-A(k). T cell responses, evident as early as 1 mo of age, depended on MHC class II binding motifs and were inhibited by anti-MHC class II Abs. Thus, although immunization can evoke peripheral self-reactive T cells in normal mice, the presence in MRL/lpr mice of spontaneous T cells reactive to certain MHC-bound self peptides suggests that these T cells actively participate in systemic autoimmunity. Peptides eluted from self MHC class II molecules may yield important clues to T cell epitopes in systemic autoimmunity.     

Effect of isotypic and allotypic variations of MHC class II molecules on staphylococcal enterotoxin presentation to murine T cells.

Staphylococcal enterotoxins (SE) are known to be potent T cell activators, stimulating +/- proliferation and lymphokine production. These toxins have recently have been termed "superantigens" because of their ability to bind directly to class II molecules forming a ligand that interacts with particular V beta gene elements within the TCR complex. This interaction between SE and MHC class II molecules plays a central role in toxin-induced mitogenesis. In the present study we have examined the effect of polymorphism on the ability of MHC class II molecules to bind and present SE. Through the use of H-2 congenic mouse strains, it was possible to look directly at haplotype differences within the MHC and their effect on SE presentation to a panel of responsive V beta-bearing T cells. The results demonstrate that toxin presentation by class II-bearing accessory cells to murine T cells is greatly affected by polymorphisms within the H-2 complex. Toxin-pulsed accessory cells obtained from mice of an H-2k and H-2u haplotype were found to be less efficient in activating a variety of T cell clones and hybridomas. However, one T cell clone responded similarly to the enterotoxins presented on all H-2 haplotypes, suggesting that differences in responses of T cells are not simply a function of the degree of binding of these toxins to various class II molecules. Neutralization analysis with monoclonal anti-class II antibodies demonstrates that both I-A and I-E molecules play a significant role in SEA and SEB presentation to murine T cells. These results suggest that the differential activation of T cells by a particular enterotoxin may reflect a difference in recognition of an SE:class II ligand by a surface T cell receptor complex.     

Efficient delivery of T cell epitopes to APC by use of MHC class II-specific Troybodies

A major objective in vaccine development is the design of reagents that give strong, specific T cell responses. We have constructed a series of rAb with specificity for MHC class II (I-E). Each has one of four different class II-restricted T cell epitopes genetically introduced into the first C domain of the H chain. These four epitopes are: 91-101 lambda2(315), which is presented by I-E(d); 110-120 hemagglutinin (I-E(d)); 323-339 OVA (I-A(d)); and 46-61 hen egg lysozyme (I-A(k)). We denote such APC-specific, epitope-containing Ab "Troybodies." When mixed with APC, all four class II-specific Troybodies were approximately 1,000 times more efficient at inducing specific T cell activation in vitro compared with nontargeting peptide Ab. Furthermore, they were 1,000-10,000 times more efficient than synthetic peptide or native protein. Conventional intracellular processing of the Troybodies was required to load the epitopes onto MHC class II. Different types of professional APC, such as purified B cells, dendritic cells, and macrophages, were equally efficient at processing and presenting the Troybodies. In vivo, class II-specific Troybodies were at least 100 times more efficient at targeting APC and activating TCR-transgenic T cells than were the nontargeting peptide Ab. Furthermore, they were 100-100,000 times more efficient than synthetic peptide or native protein. The study shows that class II-specific Troybodies can deliver a variety of T cell epitopes to professional APC for efficient presentation, in vitro as well as in vivo. Thus, Troybodies may be useful as tools in vaccine development.     

Cutting edge: H-2DM is responsible for the large differences in presentation among peptides selected by I-Ak during antigen processing

We quantitated the amounts of peptides from hen egg-white lysozyme presented by I-A(k) molecules in APC lines. The large chemical gradient of presentation of the four hen egg-white lysozyme epitopes observed in cell lines expressing HLA-DM or H-2DM (referred to in this study as DM) was significantly diminished in the T2.A(k) line lacking DM. Differences in levels of presentation between wild-type and DM-deficient APC were observed for all four epitopes, but differences were most evident for the highest affinity epitope. As a result of these quantitative differences in display, presentation of all four epitopes to T cells was impaired in the line lacking DM. The binding affinity of the pool of naturally processed peptides from DM-expressing lines was higher than that from the DM-deficient line. Thus, using a direct biochemical approach in APC, we demonstrate that DM influences the selection of peptides bound to MHC class II by favoring high affinity peptides.     

On the self-referential nature of naive MHC class II-restricted T cells

The use of mutant mice expressing a normal MHC class II molecule surface level but a severely restricted self-peptide diversity (H-2Malpha(-/-)) previously revealed that T cells carrying the Ealpha(52-68)-I-A(b) complex-specific 1H3.1 TCR rely on self-peptide(s) recognition for both their peripheral persistence in irradiated hosts and their intrathymic positive selection. Here, we identify Ealpha(52-68) structurally related self-peptide(s) as a major contributor to in vivo positive selection of 1H3.1 TCR-transgenic thymocytes in I-A(b+)/I-Ealpha(-) mice. This is demonstrated by the drastic and specific reduction of the TCR high thymocyte population in 1H3.1 TCR-transgenic (Tg) mice treated with the Ealpha(52-68)-I-A(b) complex-specific Y-Ae mAb. Self-peptide(s) recognition is also driving the maturation of T cells carrying a distinct MHC class II-restricted specificity (the Ealpha(6) alphass TCR), since positive selection was also deficient in Ealpha(6) TCR Tg H-2Malpha(-/-) thymi. Such a requirement for recognition of self-determinants was mirrored in the periphery; Ealpha(6) TCR Tg naive T cells showed an impaired persistence in both H-2Malpha(-/-) and I-A(b)ss(-/-) irradiated hosts, whereas they persisted and slowly cycled in wild-type recipients. This moderate self-peptide(s)-dependent proliferation was associated with a surface phenotype intermediate between those of naive and activated/memory T cells; CD44 expression was up-regulated, but surface expression of other markers such as CD62L remained unaltered. Collectively, these observations indicate that maturation and maintenance of naive MHC class II-restricted T cells are self-oriented processes.     

Structure, regulatory polymorphisms, and allelic hypervariability regions in murine I-A alpha

Class II major histocompatibility complex (MHC) molecules, the Ia antigens, are intimately involved in regulating the intensity and specificity of the cellular and humoral responses to T cell-dependent antigens. One approach to understanding the mechanism of this regulation is to analyze the structure and allelic polymorphism of Ia molecules. In addition there are regulatory polymorphisms in the expression of the I-E alpha and I-E beta class II MHC polypeptide chains. Analysis of the cDNA sequence indicates that I-A and I-E alpha chains are similar with short stretches of homology and other regions of nonhomology. Analysis of Northern blots of mRNA indicates that at least three separate types of regulatory polymorphisms result in failure of expression of I-E alpha. Comparison of allelic sequences of six alleles of the I-A alpha chain shows that almost all of the allelic polymorphism is in the first domain and that within the first domain it is clustered in three allelic hypervariable regions within the first domain of I-A alpha. The structural and functional implications of these findings are discussed.     

First signature of islet beta-cell-derived naturally processed peptides selected by diabetogenic class II MHC molecules

The diversity of Ags targeted by T cells in autoimmune diabetes is unknown. In this study, we identify and characterize a limited number of naturally processed peptides from pancreatic islet beta-cells selected by diabetogenic I-A(g7) molecules of NOD mice. We used insulinomas transfected with the CIITA transactivator, which resulted in their expression of class II histocompatibility molecules and activation of diabetogenic CD4 T cells. Peptides bound to I-A(g7) were isolated and examined by mass spectrometry: some peptides derived from proteins present in secretory granules of endocrine cells, and a number were shared with cells of neuronal lineage. All proteins to which peptides were identified were expressed in beta cells from normal islets. Peptides bound to I-A(g7) molecules contained the favorable binding motif characterized by acidic amino acids at the P9 position. The draining pancreatic lymph nodes of prediabetic NOD mice contained CD4 T cells that recognized three different natural peptides. Furthermore, four different peptides elicited CD4 T cells, substantiating the presence of such self-reactive T cells. The overall strategy of identifying natural peptides from islet beta-cells opens up new avenues to evaluate the repertoire of self-reactive T cells and its role in onset of diabetes.     

MHC class II A alpha and E alpha molecules determine the clonal deletion of V beta 6+ T cells. Studies with recombinant and transgenic mice

Interactions between MHC class II genes and minor lymphocyte stimulating (Mls) associated products are responsible for clonally deleting self-reactive T cells in mice. Here we demonstrate the role of the intact I-A and I-E molecules as well as the individual A alpha and E alpha chains in the deletion of cells bearing the V beta 6 TCR. DBA/1 (H-2q, Mls-1a) mice were crossed with various inbred congenic, recombinant, and transgenic strains and the F1's were screened for V beta 6 expression. All I-E+ strains were fully permissive in deleting V beta 6+ T cells. I-E- strains expressing I-A b,f,s,k,p permitted only partial deletion, while I-Aq strains showed no deletion. Recombinant I-Aq and I-Af strains which expressed E kappa alpha chain in the absence of E beta chain showed a decrease in V beta 6+ T cells as compared to their H-2q and H-2f counterparts. Furthermore, transgenic mice expressing E kappa alpha Aq beta gene in an H-2q haplotype (E kappa alpha Aq beta?) gave similar results to that of the recombinants in deleting V beta 6 T-cells. The role of the 1-A molecule was also shown by the partial deletion of V beta 6+ T cells in H-2q mice expressing transgenic I-Ak molecules. These results demonstrate that the E alpha chain is important in the deletion of V beta 6 T-cells in Mls-1a mice. The role of A alpha chain is also implied by the permissiveness of E kappa alpha Aq beta but not Aq alpha Aq beta molecules in the deletion of V beta 6+ T cells.     

Expression of I-A and I-E,C region-coded Ia antigens on functional B cell subpopulations.

Ia antigens from specific subregions have been examined on functional B cell populations. Expression of both I-A and I-E,C region antigens was demonstrated on cells required for both lipopolysaccharide mitogenesis and polyclonal activation. Similar I-A and I-E,C subregion expression was found on cells required for response to the T-independent antigen, polyvinylpyrrolidone. TNP-specific IgM and hen egg lysozyme-specific IgG plaque-forming cells also express I-A and I-E,C region antigens. No evidence was found for an Ia- population responsive in the systems tested. Further, no evidence of preferential expression of I-A or I-E,C region antigens was observed in any system examined. Therefore, it appears that B cells express both I-A and I-E,C region-coded Ia antigens.     

Selective expression of class II E alpha d gene in transgenic mice

Class II genes of the MHC must be expressed by APC for activation of CD4+ T cells and efficient delivery of T cell help to B lymphocytes. Class II genes have restricted tissue expression and are under complex regulation. By using various deletion constructs of the class II E alpha d gene in transgenic mice we have mapped different 5' flanking regions which control E alpha d gene expression in distinct cell types. We demonstrate dissociate expression of E alpha d within the macrophage lineage as well as within the B cell lineage, and present evidence for a repressive element operative in B cells and macrophages. We describe the generation of novel transgenic lines with limited constitutive and inducible E alpha mRNA and I-E protein.