T cell receptor (TCR) usage determines disease susceptibility in experimental autoimmune encephalomyelitis: studies with TCR V beta 8.2 transgenic mice

Experimental allergic encephalomyelitis (EAE) is an autoimmune disease that can be induced in laboratory animals by immunization with the major myelin proteins, myelin basic protein (MBP) and proteolipid protein (PLP). We analyzed the role of the T cell receptor (TCR) repertoire in susceptibility to EAE induced by these two autoantigens. Autoreactive T cells induced after immunization with MBP use a limited set of TCR. In contrast, we demonstrate that T cell clones that recognize the encephalitogenic PLP epitope (PLP 139-151) use diverse TCR genes. When the TCR repertoire is limited by introduction of a novel rearranged TCR V beta 8.2 chain in transgenic SJL mice, EAE could be induced in the transgenic mice by immunization with the encephalitogenic epitopes of PLP, but not with the encephalitogenic epitope of MBP. Thus, skewing the TCR repertoire affects the susceptibility to EAE by immunization with MBP but not with PLP. These data demonstrate the biological consequences of the usage of a more diverse T cell repertoire in the development of an autoimmune disease.     

CD8 surface levels alter the fate of alpha/beta T cell receptor-expressing thymocytes in transgenic mice.

The mature T cell receptor (TCR) repertoire is established on the basis of discriminative events involving binding of the TCR alpha and beta chains and CD4 or CD8 on immature thymocytes to major histocompatibility complex (MHC)/self-peptide complexes expressed in the thymus. To ask whether the strength of the interaction between a CD8/TCR complex and a MHC/self-peptide ligand plays a pivotal role in deciding the fate of a maturing thymocyte, we generated lines of transgenic mice that express distinct and elevated levels of CD8 alpha, approximately 2, 3, and 6-10 times. These lines were then crossed to a transgenic line expressing the class I-restricted TCR, 2C. We found that thymocytes expressing the 2C TCR in combination with the highest levels of CD8 were deleted on the H-2 Kb background that is normally positively selecting for the 2C TCR. In contrast, thymocytes coexpressing the 2C TCR and moderately elevated levels of CD8 were selected for maturation. These results demonstrate directly that CD8 levels can affect the developmental fate of a maturing thymocyte and argue in support of an affinity model for thymocyte selection.     

Structure of the T cell antigen receptor (TCR): two CD3 epsilon subunits in a functional TCR/CD3 complex

Transgenic mice carrying and expressing the human CD3 epsilon gene incorporate the corresponding protein product into T cell receptor (TCR)/CD3 complexes on thymocyte and T cell surfaces. The chimeric antigen receptors allow normal T cell development and selection of repertoires in vivo and are able to transduce activation signals in vitro. We have exploited the ability to distinguish mouse (m) and human (h)CD3 epsilon chains to analyze the stoichiometry of CD3 epsilon in transgenic mouse TCRs. Immunoprecipitation and fluorescence resonance energy transfer experiments demonstrate that such TCRs can contain both h- and mCD3 epsilon chains, implying that more than one CD3 epsilon subunit occurs per TCR. Antigen comodulation studies are consistent with a stochastic use of h- or mCD3 epsilon during receptor assembly, and further suggest a structure for the TCR/CD3 complex with two CD3 epsilon chains. The determination of CD3 epsilon subunit stoichiometry, together with existing biochemical data, allows the generation of a minimal model for the structure of the TCR and illustrates the potential value of the transgenic approach to the analysis of complex receptors.     

T cell receptor (TCR)-mediated repertoire selection and loss of TCR vbeta diversity during the initiation of a CD4(+) T cell response in vivo

We recently described a novel way to isolate populations of antigen-reactive CD4(+) T cells with a wide range of reactivity to a specific antigen, using immunization with a fixed dose of nominal antigen and FACS((R)) sorting by CD4(high) expression. Phenotypic, FACS((R)), functional, antibody inhibition, and major histocompatibility complex-peptide tetramer analyses, as well as T cell receptor Vbeta sequence analyses, of the antigen-specific CD4(high) T cell populations demonstrated that a diverse sperm whale myoglobin 110-121-reactive CD4(+) T cell repertoire was activated at the beginning (day 3 after immunization) of the immune response. Within 6 d of immunization, lower affinity clones were lost from the responding population, leaving an expanded population of oligoclonal, intermediate affinity (and residual high affinity) T cells. This T cell subset persisted for at least 4 wk after immunization and dominated the secondary immune response. These data provide evidence that CD4(+) T cell repertoire selection occurs early in the immune response in vivo and suggest that persistence and expansion of a population of oligoclonal, intermediate affinity T cells is involved in CD4(+) T cell memory.     

CD4(+) T cells from glutamic acid decarboxylase (GAD)65-specific T cell receptor transgenic mice are not diabetogenic and can delay diabetes transfer

Glutamic acid decarboxylase (GAD)65 is an early and important antigen in both human diabetes mellitus and the nonobese diabetic (NOD) mouse. However, the exact role of GAD65-specific T cells in diabetes pathogenesis is unclear. T cell responses to GAD65 occur early in diabetes pathogenesis, yet only one GAD65-specific T cell clone of many identified can transfer diabetes. We have generated transgenic mice on the NOD background expressing a T cell receptor (TCR)-specific for peptide epitope 286-300 (p286) of GAD65. These mice have GAD65-specific CD4(+) T cells, as shown by staining with an I-A(g7)(p286) tetramer reagent. Lymphocytes from these TCR transgenic mice proliferate and make interferon gamma, interleukin (IL)-2, tumor necrosis factor (TNF)-alpha, and IL-10 when stimulated in vitro with GAD65 peptide 286-300, yet these TCR transgenic animals do not spontaneously develop diabetes, and insulitis is virtually undetectable. Furthermore, in vitro activated CD4 T cells from GAD 286 TCR transgenic mice express higher levels of CTL-associated antigen (CTLA)-4 than nontransgenic littermates. CD4(+) T cells, or p286-tetramer(+)CD4(+) Tcells, from GAD65 286-300-specific TCR transgenic mice delay diabetes induced in NOD.scid mice by diabetic NOD spleen cells. This data suggests that GAD65 peptide 286-300-specific T cells have disease protective capacity and are not pathogenic.     

Long-term phenotypic, functional and genetic stability of cancer-specific T-cell receptor (TCR) alphabeta genes transduced to CD8+ T cells

In adoptive T-cell transfer as an intervention for malignant diseases, retroviral transfer of T-cell receptor (TCR) genes derived from CD8(+) cytotoxic T-lymphocyte (CTL) clones provides an opportunity to generate a large number of T cells with the same antigen specificity. We cloned the TCR-alphabeta genes from a human leukocyte antigen (HLA)-A(*)2402-restricted CTL clone specific for MAGE-A4(143-151). The TCR-alphabeta genes were transduced to 99.2% of non-TCR expressing SupT1, a human T-cell line, and to 12.7-32.6% of polyclonally activated CD8(+) T cells by retroviral transduction. As expected, TCR-alphabeta gene-modified CD8(+) T cells showed cytotoxic activity and interferon-gamma production in response to peptide-loaded T2-A(*)2402 and tumor cell lines expressing both MAGE-A4 and HLA-A(*)2402. A total of 24 clones were established from TCR-alphabeta gene-transduced peripheral blood mononuclear cells and all clones were functional on a transduced TCR-dependent manner. Four clones were kept in culture over 6 months for analyses in detail. The transduced TCR-alphabeta genes were stably maintained phenotypically, functionally and genetically. Our results indicate that TCR-transduced alphabeta T cells by retroviral transduction represent an efficient and promising strategy for adoptive T-cell transfer for long term.     

Stoichiometry of the T cell antigen receptor (TCR) complex: each TCR/CD3 complex contains one TCR alpha, one TCR beta, and two CD3 epsilon chains

The stoichiometry of the subunits that comprise the T cell antigen receptor (TCR) complex is not completely known. In particular, it is uncertain whether TCR alpha and TCR beta proteins are present in the TCR complex as one or multiple heterodimeric pairs. In this study we have used mice transgenic for two different TCR alpha and two different TCR beta proteins to determine the number of TCR alpha and TCR beta chains in a single TCR complex. Individual thymocytes and splenic T cells from double TCR transgenic mice simultaneously expressed all four transgenic TCR proteins on their surfaces. Because the individual TCR alpha and individual TCR beta proteins were biochemically distinguishable, we were able to examine association among the transgenic TCR products. We found that each TCR alpha chain paired with each TCR beta chain, but that each TCR complex contained only one TCR alpha and one TCR beta protein. Furthermore, quantitative immunofluorescence revealed that T cells expressed twice as many CD3 epsilon as TCR beta proteins. These findings demonstrate that there are precisely one TCR alpha, one TCR beta, and two CD3 epsilon chains in each TCR/CD3 complex expressed on the surfaces of both thymocytes and mature T cells.     

CD8 T cell memory in B cell-deficient mice

Antigen presentation by B cells and persistence of antigen-antibody complexes on follicular dendritic cells (FDC) have been implicated in sustaining T cell memory. In this study we have examined the role of B cells and antibody in the generation and maintenance of CD8+ cytotoxic T lymphocyte (CTL) memory. To address this issue we compared CTL responses to lymphocytic choriomeningitis virus (LCMV) in normal (+/+) versus B cell-deficient mice. The CTL response to acute LCMV infection can be broken down into three distinct phases: (a) the initial phase (days 3-8 after infection) of antigen-driven expansion of virus- specific CD8+ T cells and the development of effector CTL (i.e., direct ex vivo killers); (b) a phase of death (between days 10 and 30 after infection) during which >95% of the virus-specific CTL die and the direct effector activity subsides; and (c) the phase of long-term memory (after day 30) that is characterized by a stable pool of memory CTL that persist for the life span of the animal. The role of B cells in each of these three phases of the CTL response was analyzed. We found that B cells were not required for the expansion and activation of virus-specific CTL. The kinetics and magnitude of the effector CTL response, as measured by direct killing of infected targets by ex vivo isolated splenocytes, was identical in B cell-deficient and +/+ mice. Also, the expansion of CD8+ T cells was not affected by the absence of B cells and/or antibody; in both groups of mice there was an approximately 10,000-fold increase in the number of LCMV-specific CTL and a greater than 10-fold increase in the total number of activated (CD44hi) CD8+ T cells during the first week after virus infection. Although no differences were seen during the "expansion" phase, we found that the "death" phase was more pronounced in B cell-deficient mice. However, this increased cell death was not selective for LCMV- specific CTL, and during this period the total number of CD8+ T cells also dropped substantially more in B cell-deficient mice. As a result of this, the absolute numbers of LCMV-specific CTL were lower in B cell- deficient mice but the frequencies were comparable in both groups of mice. More significantly, the memory phase of the CTL response was not affected by the absence of B cells and a stable number of LCMV-specific CTL persisted in B cell-deficient mice for up to 6 mo. Upon reinfection, B cell-deficient mice that had resolved an acute LCMV infection were able to make accelerated CTL responses in vivo and eliminated virus more efficiently than naive B cell-deficient mice. Thus, CTL memory, as assessed by frequency of virus-specific CTL or protective immunity, does not decline in the absence of B cells. Taken together, these results show that neither B cells nor antigen-antibody complexes are essential for the maintenance of CD8+ CTL memory.     

T cells with gamma/delta T cell receptors (TCR) of intestinal type are preferentially expanded in TCR-alpha-deficient lpr mice

Fas-mediated apoptosis is essential for activation-induced cell death of alpha/beta T cells, but it is not clear what role, if any, it plays in regulating other components of the immune system. To study the role of Fas in gamma/delta T cell development, Fas-deficient lpr mice were bred with T cell receptor alpha gene-ablated (TCR-alpha-/-) mice to generate mice deficient in one or both genes. The TCR-alpha-/-, lpr/lpr mice had a nearly 10-fold increase in total lymph node cell (LNC) number compared with Fas-intact TCR-alpha-/- mice, because of expansion of TCR-gamma/delta+ and TCR-beta+ cells. In Fas-intact TCR-alpha-/- mice, approximately one third of the LNCs expressed TCR-gamma/delta. These were evenly divided between the CD4-, CD8-alpha+ and the CD4-, CD8- subsets, and rarely expressed the B220 epitope of CD45. In contrast, in TCR-alpha-/-, lpr/lpr mice, TCR-gamma/delta+ cells comprised half of the LNCs and were primarily CD4-, CD8-, and B220+. Moreover, Fas deficiency in TCR-alpha-/- mice caused a preferential expansion of gamma/delta T cells expressing variable region genes characteristic of intestinal intraepithelial lymphocytes. These results demonstrate a role for Fas in regulating the gamma/delta T cell contribution to peripheral lymph nodes. This mechanism may be most important in limiting the access of activated intestinal intraepithelial lymphocytes to the peripheral lymphoid system.     

Analysis of T cell antigen receptor (TCR) expression by human peripheral blood CD4-8- alpha/beta T cells demonstrates preferential use of several V beta genes and an invariant TCR alpha chain

CD4-CD8- (double negative [DN]) alpha/beta T cells are a largely uncharacterized subpopulation of unknown function. To investigate whether these cells are selected to recognize particular antigens or antigen-presenting molecules, DN alpha/beta T cells were purified from the peripheral blood of five normal donors and their T cell receptor (TCR) alpha and beta chains were examined. Random cloning of TCR alpha chains by single-sided polymerase chain reaction (PCR) amplification identified an invariant rearrangement between V alpha 24 and J alpha Q, with no N region diversity, which was expressed preferentially by DN alpha/beta T cells from all donors. Random cloning also identified a precise V alpha 7.2-J alpha (IGRJa14) rearrangement, with two variable amino acids encoded in the V-J junction, which was enriched in the DN alpha/beta T cell preparations from some, but not all, donors. Analysis of TCR beta chains by quantitative PCR amplification demonstrated that the expression of four V beta gene families, V beta 2, 8, 11, and 13, was markedly increased in these DN alpha/beta T cell preparations. The expression of particular TCRs by DN alpha/beta T cells from multiple donors indicates that these cells, or at least a subpopulation of cells with this phenotype, recognize a limited spectrum of antigens and suggests that they may use nonpolymorphic antigen-presenting molecules.     

Expression of two alpha chains on the surface of T cells in T cell receptor transgenic mice

CD8+ T cells taken directly from mice expressing a Kb-specific T cell receptor (TCR) transgene expressed the transgenic TCR in a bimodal profile as detected by flow cytometric analysis using a clonotype- specific monoclonal antibody. Those cells expressing the lower density of the transgenic TCR expressed the transgenic beta chain and two different alpha chains on their surface. One alpha chain was the product of the alpha transgene, whereas the other was derived by endogenous rearrangement. This report provides the first demonstration that T cells isolated directly from mice may express two different TCR clonotypes on their surface. The potential consequences of this finding for studies using TCR transgenic mice and for the induction of autoimmunity are discussed.     

TCR stimulation with modified anti-CD3 mAb expands CD8+ T cell population and induces CD8+CD25+ Tregs

Modified anti-CD3 mAbs are emerging as a possible means of inducing immunologic tolerance in settings including transplantation and autoimmunity such as in type 1 diabetes. In a trial of a modified anti-CD3 mAb [hOKT3gamma1(Ala-Ala)] in patients with type 1 diabetes, we identified clinical responders by an increase in the number of peripheral blood CD8+ cells following treatment with the mAb. Here we show that the anti-CD3 mAb caused activation of CD8+ T cells that was similar in vitro and in vivo and induced regulatory CD8+CD25+ T cells. These cells inhibited the responses of CD4+ cells to the mAb itself and to antigen. The regulatory CD8+CD25+ cells were CTLA4 and Foxp3 and required contact for inhibition. Foxp3 was also induced on CD8+ T cells in patients during mAb treatment, which suggests a potential mechanism of the anti-CD3 mAb immune modulatory effects involving induction of a subset of regulatory CD8+ T cells.     

Developmental arrest of NK1.1+ T cell antigen receptor (TCR)- alpha/beta+ T cells and expansion of NK1.1+ TCR-gamma/delta+ T cell development in CD3 zeta-deficient mice

The relationship between the structure of the T cell antigen receptor (TCR)-CD3 complex and development of NK1.1+ T cells was investigated. The TCR complex of freshly isolated NK1.1+ TCR-alpha/beta+ thymocytes contained CD3 zeta homodimers and CD zeta-FcR gamma heterodimers, whereas that of the majority of NK1.1- T cells did not contain FcR gamma. The function of CD3 zeta and FcR gamma in the development of NK1.1+ T cells was determined by analyzing CD3 zeta- and FcR gamma- deficient mice. The NK1.1+ T cells from wild-type and CD3 zeta- deficient mice had equal levels of CD3 expression. However, the development of NK1.1+ TCR-alpha/beta+ T cells was almost completely disrupted in thymus and spleen in CD3 zeta-deficient mice, whereas no alteration was observed in FcR gamma-deficient mice. In contrast, the number of novel NK1.1+ TCR-gamma/delta+ thymocytes expressing a surface phenotype similar to NK1.1+ TCR-alpha/beta+ thymocytes increased approximately six times in CD3 zeta-deficient mice. These findings establish the distinct roles of the CD3 zeta chain in the development of the following different thymic T cell compartments: NK1.1- TCR+, NK1.1+ TCR-alpha/beta+, and NK1.1+ TCR-gamma/delta+ thymocytes, which cannot be replaced by CD3 eta or FcR gamma chains.     

CD8 and β2-microglobulin-free MHC class I molecules in T cell immunoregulationMHC class I molecules in T cell immunoregulation

Summary Intracellular assembly of MHC class I heavy chains with β₂-microglobulin occurs prior to the expression of the antigen-presenting complex on the cell surface. The association of β₂-microglobulin with newly synthesized class I heavy chains is thought to be a strict prerequisite for their transport to the cell surface. However, MHC class I molecules not associated with β₂-microglobulin (β₂-microglobulin-free class I heavy chains) have been detected on the surface of activated lymphoid cells. These molecules have different conformations. Therefore, their interactions with other membrane proteins and biological functions may be different from those assigned to β₂-microglobulin-associated MHC class I molecules. The two forms of MHC class I molecules on the surface of activated cells can self-associated and also form complexes with distinct proteins. Upon interaction with the appropriate ligands these molecular complexes transduce signals regulating cell activation. The ligand for β₂-microglobulin-free class I heavy chains appears to be soluble CD8. A model is presented describing a novel mechanism of immunoregulation mediated by both soluble and membrane-bound forms of CD8 and β₂-microglobulin-free class I heavy chains.     

The presence of interleukin 4 during in vitro priming determines the lymphokine-producing potential of CD4+ T cells from T cell receptor transgenic mice

To study the factors that determine whether CD4+ T cells produce interleukin 4 (IL-4) or interferon gamma (IFN-gamma) upon stimulation we used a system allowing naive T cells to be primed in vitro by specific antigen. Dense CD4+ T cells were purified from mice that expressed transgenes encoding a T cell receptor specific for pigeon cytochrome C peptide 88-104 in association with I-Ek. These T cells produced very limited amounts of IL-4 and IFN-gamma upon immediate challenge with 88-104 and antigen-presenting cells (APC). However, after an initial "priming" culture in which they were incubated for 4 d in the presence of 88-104, APC, and 1,000 U/ml IL-4, the T cells acquired the capacity to produce substantial amounts of IL-4 upon rechallenge but made very little IFN-gamma. Cells primed in the absence of IL-4 produced IFN-gamma upon rechallenge but virtually no IL-4. The inhibitory effect of IL-4 on IFN-gamma production did not appear to be mediated by the induction of IL-10 production since IL-10 addition to initial cultures did not suppress priming for IFN-gamma production, nor did anti-IL-10 block the inhibitory effect of IL-4. IFN-gamma itself did not increase priming for IFN-gamma production, nor did anti-IFN-gamma reduce such priming. IFN-gamma did, however, diminish priming for IL-4 production when limiting amounts of IL-4 (100 U/ml) were used in the initial culture. The dominant effect of IL-4 in determining the lymphokine-producing phenotype of primed cells was observed with dendritic cells (DC), activated B cells, and I-Ek-transfected fibroblasts as APC. However, the different APC did vary in their potency, with DC being superior to activated B cells, which were superior to transfected fibroblasts.     

CD8beta endows CD8 with efficient coreceptor function by coupling T cell receptor/CD3 to raft-associated CD8/p56(lck) complexes.

The extraordinary sensitivity of CD8+ T cells to recognize antigen impinges to a large extent on the coreceptor CD8. While several studies have shown that the CD8beta chain endows CD8 with efficient coreceptor function, the molecular basis for this is enigmatic. Here we report that cell-associated CD8alphabeta, but not CD8alphaalpha or soluble CD8alphabeta, substantially increases the avidity of T cell receptor (TCR)-ligand binding. To elucidate how the cytoplasmic and transmembrane portions of CD8beta endow CD8 with efficient coreceptor function, we examined T1.4 T cell hybridomas transfected with various CD8beta constructs. T1.4 hybridomas recognize a photoreactive Plasmodium berghei circumsporozoite (PbCS) peptide derivative (PbCS (4-azidobezoic acid [ABA])) in the context of H-2K(d), and permit assessment of TCR-ligand binding by TCR photoaffinity labeling. We find that the cytoplasmic portion of CD8beta, mainly due to its palmitoylation, mediates partitioning of CD8 in lipid rafts, where it efficiently associates with p56(lck). In addition, the cytoplasmic portion of CD8beta mediates constitutive association of CD8 with TCR/CD3. The resulting TCR-CD8 adducts exhibit high affinity for major histocompatibility complex (MHC)-peptide. Importantly, because CD8alphabeta partitions in rafts, its interaction with TCR/CD3 promotes raft association of TCR/CD3. Engagement of these TCR/CD3-CD8/lck adducts by multimeric MHC-peptide induces activation of p56(lck) in rafts, which in turn phosphorylates CD3 and initiates T cell activation.     

Predominant role of T cell receptor (TCR)-alpha chain in forming preimmune TCR repertoire revealed by clonal TCR reconstitution system

The CDR3 regions of T cell receptor (TCR)-alpha and -beta chains play central roles in the recognition of antigen (Ag)-MHC complex. TCR repertoire is created on the basis of Ag recognition specificity by CDR3s. To analyze the potential spectrum of TCR-alpha and -beta to exhibit Ag specificity and generate TCR repertoire, we established hundreds of TCR transfectants bearing a single TCR-alpha or -beta chain derived from a cytotoxic T cell (CTL) clone, RT-1, specific for HIVgp160 peptide, and randomly picked up TCR-beta or -alpha chains. Surprisingly, one-third of such TCR-beta containing random CDR3 beta from naive T cells of normal mice could reconstitute the antigen-reactive TCR coupling with RT-1 TCR-alpha. A similar dominant function of TCR-alpha in forming Ag-specific TCR, though low-frequency, was obtained for lymphocytic choriomeningitis virus-specific TCR. Subsequently, we generated TCR-alpha and/or -beta transgenic (Tg) mice specific for HIVgp160 peptide, and analyzed the TCR repertoire of Ag-specific CTLs. Similar to the results from TCR reconstitution, TCR-alpha Tg generated CTLs with heterogeneous TCR-beta, whereas TCR-beta Tg-induced CTLs bearing a single TCR-alpha. These findings of Ag recognition with minimum involvement of CDR3 beta expand our understanding regarding the flexibility of the spectrum of TCR and suggest a predominant role of TCR-alpha chain in determining the preimmune repertoire of Ag-specific TCR.     

The in vivo cytotoxic activity of CD8+ T cell clones correlates with their levels of expression of adhesion molecules

CD8+ T cell clones specific for a defined epitope present in the circumsporozoite protein of Plasmodium yoelii display striking differences in their in vivo antiplasmodial activity. The adoptive transfer of certain clones (YA23 and YA26) into naive mice inhibits by 90% or more the development of liver stages of malaria parasites and protects against malaria infection. The adoptive transfer of two other T cell clones (YB8 and YA15) results, respectively, in partial or no inhibitory activity on parasite development. We found that "protective" and "nonprotective" cytotoxic T lymphocyte (CTL) clones do not differ in their fine epitope specificity and display similar levels of lysis and DNA degradation of target cells in vitro. Their pattern of production of lymphokines and granule-associated proteins also failed to correlate with their in vivo antiplasmodial activity. Histological studies combined with autoradiography showed that, upon adoptive transfer, only T cells from the protective CTL clones are capable of "associating" with a significant percentage of parasitized hepatocytes. Fluorescence-activated cell sorter analysis of surface molecules revealed pronounced differences in the levels of CD44 and VLA-4 expression by the different clones, correlating closely with their in vivo protective activity. The correlation between in vivo antiparasite activity and the expression of CD44 was further corroborated by the results of sorting, from the partially protective YB8 clone, two sub-populations expressing high and low levels of CD44. These were protective and nonprotective, respectively. The clones also differed in their adhesive properties. Cross-linking of CD44, using specific antibodies, induced LFA-1-mediated homotypic aggregation of protective clones, while nonprotective cells failed to aggregate.     

The clone size of peripheral CD8 T cells is regulated by TCR promiscuity

Positive selection in the thymus and peripheral T cell survival depend on T cell receptor (TCR)-major histocompatibility complex (MHC) interactions, but it is not yet clear if both events follow exactly the same rules. We studied peripheral T cell survival and clone sizes in conditions of progressive reduction of restricting MHC-bearing cells or progressive ablation of different MHC molecules. Different CD8(+) T cell clones/polyclonal populations showed different survival and/or lymphopenia-driven proliferation requirements. We could correlate clone sizes to the capacity of each TCR to interact with different types of MHC complexes. Thus, although repertoire selection in the thymus is mainly conditioned by the affinity of TCR-MHC interactions, peripheral selection is determined by TCR cross-reactivity to environmental ligands.