The role of adenosine receptor engagement in murine fetal thymocyte development.

The role of adenosine receptor engagement in murine T-cell development was evaluated by culturing day 15-16 fetal thymic lobes in the presence of various concentrations of the adenosine receptor agonist 5'-(N-ethyl)-carboxamidoadenosine (NECA) or the adenosine receptor antagonist 8-phenyl-theophylline (8-PT) using the fetal thymic organ culture (FTOC) system. Before and 8 days after culture, thymocytes were isolated, counted, and analyzed for the expression of CD4 and CD8 T-cell differentiation molecules. Analysis of fresh thymocytes prior to culture showed that the majority of cells were of the CD4 single-positive or CD4+ CD8+ immature phenotype. Eight days after culture with media alone, 44% of cells were CD4+ and 23% were CD8+, and the number of viable thymocytes had increased from 1.7 x 10(5) to 2.2 x 10(5) cells per thymic lobe. A dose-dependent inhibition of maturation was observed in cultures with 8-PT, with greater than 85% inhibition at 50 microM. The double-positive thymocyte subset was most severely depleted. The number of cells obtained from cultures with NECA was also reduced, with about 65% inhibition at 50 microM, especially the CD8+ subset that was most severely affected. These results suggest that adenosine receptor engagement is required for normal T-cell differentiation and that adenosine receptor agonists and antagonists have distinct effects on thymocyte differentiation. An understanding of the cell-type-specific and developmental expression of adenosine receptors will help elucidate the mechanisms by which adenosine receptor engagement influences T-cell development.     

T cell receptor-negative thymocytes from SCID mice can be induced to enter the CD4/CD8 differentiation pathway

In order to investigate the role of T cell receptor (TcR) expression in thymocyte maturation, we have analyzed thymocytes from C.B-17/SCID mice, which are unable to productively rearrange their antigen receptor genes and fail to express TcR. Despite this defect, SCID thymocytes are functional as they produce lymphokines and proliferate in response to a variety of stimuli. Phenotypic analysis revealed that thymocyte populations from young adult SCID mice resemble thymocyte populations from normal embryonic mice in that they are large, Thy-1.2+, CD4-, CD8-, TcR- and enriched in CD5lo, IL2R+ and Pgp1+ cells. However, other TcR- populations normally present in adult mice (i.e., CD4-CD8+ cells and CD4+CD8+ cells) are absent from the thymus of TcR- adult SCID mice. To understand the basis of the developmental arrest of TcR- SCID thymocytes at the CD4-CD8- stage of differentiation, we analyzed thymi from the occasional "leaky" SCID mouse which possesses small numbers of TcR+ thymocytes. We found that the presence of TcR+ cells within a SCID thymus was invariably associated with the presence of CD4+ and/or CD8+ SCID thymocytes. Interestingly, however, the CD4+/CD8+ SCID thymocytes were not themselves necessarily TcR+. That is, emergence of SCID thymocytes expressing CD4/CD8 was tightly linked to the presence of TcR+ cells within that SCID thymus, but the SCID thymocytes that expressed CD4/CD8 were not necessarily the same cells that expressed TcR. Finally, we found that the introduction into TcR- SCID mice of normal bone marrow cells that give rise to TcR+ cells within the SCID thymus promoted the differentiation of SCID thymocytes into CD4-CD8+ and CD4+CD8+ TcR- cells. These data indicate that TcR+ cells within the thymic milieu provide critical signals which promote entry of CD4-CD8-TcR- precursor T cells into the CD4/CD8 differentiation pathway. When applied to differentiation of normal thymocytes, these findings may imply a critical role for early appearing CD4-CD8- TcR (gamma/delta)+ cells in initiating normal thymic ontogeny.     

Hedgehog signaling regulates differentiation from double-negative to double-positive thymocyte

The hedgehog (Hh) signaling pathway is involved in the development of many tissues. Here we show that sonic hedgehog (Shh) is involved in thymocyte development. Our data suggest that termination of Hh signaling is necessary for differentiation from CD4-CD8-double-negative (DN) to CD4+CD8+ double-positive (DP) thymocyte. Shh is produced by the thymic stroma, and Patched and Smoothened (Smo), the transmembrane receptors for Shh, are expressed in DN thymocytes. A neutralizing monoclonal antibody against Shh increases differentiation of DN to DP thymocytes, and Shh protein arrests thymocyte differentiation at the CD25+ DN stage, after T cell receptor beta (TCRbeta) gene rearrangement. We show that one consequence of pre-TCR signaling is downregulation of Smo, allowing DN thymocytes to proliferate and differentiate.     

Alterations in thymocyte subpopulations in Down's syndrome (trisomy 21)

To correlate the histologically observed thymic abnormalities with the cellular immunodeficiency found in Down's syndrome (DS), thymus fragments and thymocyte suspensions from 14 noninstitutionalized DS subjects were studied. Histologic examination and immunohistologic studies using an anticluster of differentiation (CD) 1 monoclonal antibody showed a contracted cortex due to cortical thymocyte depletion. When DS unselected thymocytes were phenotyped, a significant reduction of CD3-, CD1-, CD4-, and CD8-positive cells was found as compared to controls. To evaluate if the deficient expression of these markers was due to the reduction of thymocyte subsets identifiable on the basis of their physical properties, we separated DS unselected thymocytes into 10 fractions by continuous Percoll density gradient centrifugation. DS thymuses were almost completely devoid of high density thymocytes. Since in normal thymus, these cells correspond to small CD1+, CD4+, CD8+, and 50% CD3+ cortical thymocytes, their absence may explain the unrestricted reduction of markers on DS unfractionated thymocytes. Furthermore DS thymuses appeared to be enriched in CD1+ first fraction (Fr1) low density thymocytes of the Percoll gradient. Fr1 CD1+ cells constitute the main spontaneously proliferating pool in normal human thymus. When the spontaneous proliferating activity of DS Fr1 was compared to that of the control, a significant reduction was observed. This reduction associated with the absence of high density thymocytes, with the reduction of cells expressing alpha- and beta-chains of the T cell receptor and in conclusion with the lymphocyte depletion, suggests that in DS thymuses there is a deficient expansion of immature T cells resulting in a reduction of the various thymocyte subpopulations, including the thymocyte pool able to differentiate into functionally mature T cells.     

Intrathymic T cell development and selection proceeds normally in the absence of glucocorticoid receptor signaling

Glucocorticoids are believed to play a role in T cell development and selection, although their precise function is controversial. Glucocorticoid receptor (GR)-deficient mice were used to directly investigate this problem. GR-deficient thymocytes were resistant to dexamethasone-mediated apoptosis, confirming the absence of glucocorticoid responsiveness. An absence of GR signaling had no impact on thymocyte development either in vivo or in vitro. T cell differentiation, including positive selection, was normal as assessed by normal development of CD4+CD8+, alphabetaTCR+CD4+, and alphabetaTCR+CD8+ thymocytes. Negative selection, mediated by the superantigen staphylococcal enterotoxin B (SEB), or anti-CD3/CD28, was also normal in the absence of GR signaling. In contrast to earlier reports, these data demonstrate that GR signaling is not essential for intrathymic T cell development or selection.     

The BTG/TOB family protein TIS21 regulates stage-specific proliferation of developing thymocytes

As thymocytes undergo differentiation in the thymus, they progress through distinct phases of quiescence and proliferation. Identifying cellular mechanisms that maintain thymocytes in a non-dividing state is critical to fully understand T cell development. A member of the B cell translocation gene/transducer of ErbB-2 (BTG/TOB) family of anti-proliferative proteins was identified as a key mediator of the quiescent state in peripheral anergic and unstimulated T cells. Here, we demonstrate that the BTG/TOB family member TPA-inducible sequence 21 (TIS21) is expressed in quiescent CD44+ CD25- early progenitor thymocytes and CD44- CD25+ cells prior to TCR beta-selection. However, TIS21 expression is decreased in proliferating CD25+ CD44+ progenitor thymocytes and CD25(low) CD44- beta-selected cells, suggesting that its regulated expression may enable thymocytes to remain quiescent in the absence of mitogenic signals. We addressed the role of TIS21 in regulating thymocyte stage-specific expansion by ectopically expressing TIS21 in developing thymocytes and hematopoietic progenitors. Dysregulated expression of TIS21 inhibited the expansion of thymocytes even in the presence of endogenous mitogenic signals, while thymocyte differentiation was unimpeded. These findings imply that the intracellular mechanisms regulating thymocyte differentiation and proliferation, which are induced downstream of developmental cues, function independently during early T cell development.     

CD4-CD8- thymocytes that express the T cell receptor may have previously expressed CD8

Amongst CD4-CD8- (double negative) thymocytes there is a sizeable population (variable from strain to strain) of cells expressing surface T cell receptor (TCR). These TCR+ double negatives are predominantly non-cycling, have very little precursor activity, and, unlike the TCR-CD4-CD8- thymocytes, appear not to be part of the mainstream of thymocyte development. A unique feature of this population is the biased V beta-gene region usage. In CBA mice, 60-70% of TCR+ CD4-CD8- cells express receptors that utilize V beta 8 gene products, compared with peripheral T cells from the same strain which are only 20-30% V beta 8+. This suggests that the high V beta 8 usage may be the result of some selective process. A growing body of experimental data suggests that TCR specificity selection occurs at the CD4+CD8+ stage of thymocyte development. In order to gain some insight into the previous history of the TCR+ double negatives, in particular whether or not they have previously expressed CD8 and therefore been eligible for selection, we have determined the methylation state of the CD8 gene and compared it to other thymocyte populations. We show that the TCR+ CD4-CD8- thymocytes are demethylated at some sites in the CD8 gene, consistent with previous CD8 expression. However, the demethylation pattern is distinct from that seen on typical peripheral T cells or on mature thymocytes, suggesting that the TCR+ CD4-CD8- thymocytes are not derived from mature thymocytes or peripheral T cells which have returned to the thymus and downregulated CD8 expression.(ABSTRACT TRUNCATED AT 250 WORDS)     

A novel thymic epithelial adhesion molecule

Interactions between thymocytes and thymic stromal cells are responsible for positive and negative selection of T cell, their differentiation, maturation and proliferation. The signals required for these events to occur often necessitate close contact, and indeed adhesion, between the cell types involved. The identification of specific adhesion molecules in this context, is, therefore, a vital first step in determining the nature of the signal they mediate or facilitate at a given stage of differentiation. In the present work we identify, isolate and partially characterize a ligand present on thymic medullary epithelial cells which selectively binds CD4+ CD8+ thymocytes found primarily in the thymic cortex. This adhesion molecule is composed of two non-covalently associated glycoproteins of 23 kDa and 45 kDa, respectively, both of which are needed to bind to thymocytes. The importance of the finding is that the ligand, in isolated immobilized form, binds the same thymocyte subset as the original epithelial cell line from which it was isolated. The CD4+ CD8+ thymocyte subset is the precursor of single-positive mature T cells; hence the putative biological activity of the ligand in question takes place at a pivotal stage of T cell differentiation.     

Accelerated transition from the double-positive to single-positive thymocytes in G alpha i2-deficient mice

Deletion of alpha i2 subunit of heterotrimeric G proteins induces a 2-4-fold increase in the proportions of CD4 and CD8 single-positive (SP) thymocytes as compared with wild-type littermates, but how G alpha i2 is involved in thymocyte development is unknown. To determine a role for G alpha i2 in a specific developmental stage of thymocyte differentiation, we studied the ontogeny of thymocytes in G alpha i2-deficient mice. Our data show that an accelerated transition from the double-positive (DP) to SP thymocytes, rather than impairment in thymic emigration, accounts for a high proportion of the SP thymocytes in the absence of G alpha i2. Lack of G alpha i2 greatly augmented a response of thymocytes to TCR-mediated stimulation, as evidenced by enhanced proliferation of the DP thymocytes upon ligation of the TCRs. The augmented response may be the reason behind the expedited transition from the DP to SP thymocytes in the animal. In accordance with this, effects of G alpha i2 deficiency on CD8 or CD4 SP thymocyte differentiation required engagement of the TCRs with either MHC class I or MHC class II molecule. The abnormal thymocyte development resulted in an increase in positive selection, altered usage of TCR Vbeta gene, aberrant development of CD4+ CD25+ T regulatory cells and untimely thymic involution, the contribution of which to colitis development in the animal is discussed. These findings reveal a previously unappreciated role for G alpha i2 protein in clonal selection and functionality of thymocytes.     

Notch1 regulates maturation of CD4+ and CD8+ thymocytes by modulating TCR signal strength

Notch signaling regulates cell fate decisions in multiple lineages. We demonstrate in this report that retroviral expression of activated Notch1 in mouse thymocytes abrogates differentiation of immature CD4+CD8+ thymocytes into both CD4 and CD8 mature single-positive T cells. The ability of Notch1 to inhibit T cell development was observed in vitro and in vivo with both normal and TCR transgenic thymocytes. Notch1-mediated developmental arrest was dose dependent and was associated with impaired thymocyte responses to TCR stimulation. Notch1 also inhibited TCR-mediated signaling in Jurkat T cells. These data indicate that constitutively active Notch1 abrogates CD4+ and CD8+ maturation by interfering with TCR signal strength and provide an explanation for the physiological regulation of Notch expression during thymocyte development.     

Interleukin-7 treatment promotes the differentiation pathway of T-cell-receptor-alpha beta cells selectively to the CD8+ cell lineage.

In this report we have studied the influence of interleukin-7 (IL-7) on thymocyte differentiation by evaluating the effects of IL-7 on the generation of T-cell receptor-alpha beta (TCR-alpha beta) and TCR-gamma delta thymocyte subpopulations in rat fetal thymus organ culture. IL-7 enhanced the differentiation pathway of TCR alpha beta thymocytes, first increasing the numbers of immature CD8+ cells, and later those of both CD4+ CD8+ and mature thymocytes. The kinetics of thymocyte migration out of thymic lobes was also accelerated, and the average number of mature TCR-alpha beta phi emigrants per day was increased in the presence of IL-7. Moreover, mature CD4- CD8+ thymocytes were preferentially generated after IL-7 administration. This TCR-alpha beta hi cell population was not actively dividing, indicating that IL-7-promoted thymocyte differentiation was selective to the CD8 cell lineage. Distribution of some TCR-V alpha and TCR-V beta segments among mature thymocytes was also modified in IL-7-treated thymic lobes. On the contrary, the maturation of TCR-gamma delta was not affected by IL-7 addition during the first days of culture, but their numbers sharply increased by day 6 of culture. These results were confirmed with IL-7-treated cultures for 24 hr, showing that IL-7 responsiveness was acquired by TCR-gamma delta cells late in thymus ontogeny. The present results thus indicate a key role for IL-7 in the maturation of TCR-alpha beta thymocytes and the expansion of thymic TCR-gamma delta cells.     

Delineation of chicken thymocytes by CD3-TCR complex, CD4 and CD8 antigen expression reveals phylogenically conserved and novel thymocyte subsets.

To further define the relationship between thymocyte subsets and their developmental sequence, multi-parameter flow cytometry was used to determine the distribution of the CD3-TCR complex and the accessory molecules CD4 and CD8 on chicken thymocytes. As in mammals, adult thymocytes could be subdivided into CD3-, CD3lo, and CD3hi staining populations. CD4 and CD8 distribution on such populations revealed the presence of CD3-CD4+CD8- and CD3-CD4-CD8+ thymocytes, putative precursors to CD4+CD8+ cells, detectable in the adult and at high frequency during ontogeny. Of particular interest was the existence of CD3lo expression on CD4+CD8- and CD4-CD8+, and in some instances, on CD4-CD8- thymocytes. Such phenotypes are not easily detectable in the mammalian thymus but were readily observed in both adult and embryonic chicken thymus from 16 days of embryogenesis. Further analysis of the TCR lineage of these CD3lo cells revealed that they were essentially all of the alpha beta TCR type. Mature CD3hi thymocytes were found within the CD4+CD8+ and CD4+CD8- and CD4-CD8+ subsets. Both alpha beta and gamma delta TCR lineage thymocytes were detected within all CD4- and CD8-defined subsets, thus identifying novel thymocyte subsets in the chicken thymus, namely alpha beta TCR+CD4-CD8- and gamma delta TCR+ CD4+CD8- cells. Hence, this analysis of chicken thymocytes, while confirming the phylogenically conserved nature of the thymus, has revealed novel T cell subsets, providing further insight into the complexity of mainstream thymocyte maturation pathways.     

In vitro differentiation of a CD4/CD8 double-positive equivalent thymocyte subset in adult Xenopus.

The thymus is the major site of selection and differentiation of T cells in mammals and birds. To begin to study the evolution of thymocyte differentiation, we have developed, in the frog Xenopus, an in vitro system that takes advantage of cortical thymocyte antigen (CTX), a recently discovered T cell antigen whose expression is restricted to Xenopus cortical thymocytes. Upon transient stimulation with suboptimal mitogenic concentrations of the phorbol ester phorbol myristate acetate (PMA) plus ionomycin, Xenopus thymocytes are induced to differentiate into cycling T lymphoblasts that actively synthesize and express high levels of surface MHC class I and class II molecules. This appearance of T lymphoblasts correlates with a rapid down-regulation of both surface CTX protein and CTX mRNA. A thymocyte subset with an immature phenotype (CTX+, CD8+, class II- or class II low and class I-) was characterized by depleting class II+ cells or by panning with anti-CTX mAb. This immature CTX+ thymocyte subset displays a limited proliferative capacity compared to total, class II+ or to CTX- thymocytes, and can be induced, by PMA/ionomycin, to differentiate into more mature T lymphoblasts expressing surface class II and class I molecules. These results provide the first in vitro evidence in an ectothermic vertebrate of a conserved intrathymic pathway of thymocyte differentiation. In addition, our data reveal that CTX can serve as a differentiation surface marker of a population of immature thymocytes that appears to be the equivalent of the mammalian CD4/CD8 double-positive subset.     

Interplay between IL-2 and IL-4 in human thymocyte differentiation: antagonism or agonism

The effect of recombinant interleukin 2 (IL-2) and IL-4, as well as a combination of both lymphokines on human post-natal thymocytes at different maturation stages, was analyzed by culturing highly purified pro-T cells, pre-T cells, double-negative and double-positive thymocyte subsets in the presence of IL-2 and/or IL-4. Both IL-2 and IL-4 responsiveness are developmentally regulated in human thymocytes, since IL-2 and IL-4 responses decline with increasing thymocyte differentiation, double-positive T cells displaying far less proliferation than immature thymocytes. IL-2 and IL-4 may influence pro-T cell growth in both an antagonistic and additive fashion. At low doses, IL-4 inhibits IL-2-supported growth of pro-T cells, whereas, at higher concentrations, this inhibitory effect is masked by the ability of IL-4 to stimulate pro-T cell proliferation. In contrast to peripheral lymphocytes, IL-4 does not down-regulate the expression of the IL-2 receptor light chain on thymocytes. In pro-T cell cultures, IL-2 and IL-4 favour the differentiation of distinct cell populations, namely lymphocytes displaying preferentially a TCR alpha/beta+ and CD4+CD8- phenotype versus predominantly TCR gamma/delta+ and CD4-CD8+ cells, respectively. The effect of IL-2 dominates over that of IL-4, since the composition of cultures set up in the presence of IL-2 plus IL-4 resembles that of cells cultured with IL-2 alone. In synthesis, IL-2 and IL-4 exhibit reciprocal inter-relations in human thymocyte cultures, thus supporting the notion that these lymphokines are implicated in the complex regulation of a local cytokine network.     

TCR signaling inhibits glucocorticoid-induced apoptosis in murine thymocytes depending on the stage of development

Signaling by either the TCR or glucocorticoid receptor (GR) induces apoptosis in thymocytes. Interestingly, it has been shown previously that hybridoma T cells escape apoptosis induced by either TCR or GR when both of these receptors signal simultaneously. Whether such mutual antagonism is present in primary thymocytes was the subject of the present study. Both glucocorticoids (GC) and anti-TCR/CD28 (or anti-CD3/CD28) mAb induced apoptosis in total thymocytes. When these signals were present at the same time, GC-induced apoptosis was partially inhibited by TCR/CD3 signaling. Costimulation by anti-CD28 enhanced the inhibitory effects of anti-CD3 on GC-induced apoptosis about 30-fold. However, subset analysis revealed that most cells rescued from GC-induced apoptosis were mature CD4+ and CD8+ thymocytes, and these cells were resistant to TCR/CD3-induced apoptosis in the absence of GC. Similar results were obtained with mature splenic CD4+ and CD8+ T cells. TCR/CD3 signaling alone, while inducing apoptosis in CD4+(CD8+)TCRlow thymocytes, rescued a small subset of CD4+(CD8+)TCRlow thymocytes from GC-induced apoptosis. Thus, TCR signaling increasingly reverses GC-induced apoptosis as thymocyte development progresses. As GC are infinitely present in vivo, these findings support a model wherein TCR signaling may be required to prevent GC-induced apoptosis both under basal and immune challenging conditions.     

Restoration of thymic development in an Lck(-/-) thymoma overexpressing ZAP-70

Thymic development is strictly controlled by Src and Syk family protein tyrosine kinases. The major players in this process are Lck and ZAP-70, which regulate critical differentiation steps of thymopoiesis. Notwithstanding the critical role of Lck and ZAP-70 in thymocyte development as compared to the related kinases Fyn and Syk, a partial functional redundancy between members of the same family of protein tyrosine kinases has emerged from studies on genetically manipulated mouse models. Furthermore, a close functional interplay between Lck and ZAP-70 in intracellular signaling has been shown to occur in thymocytes. Here we present the characterization of a thymoma from an Lck(-/-) mouse, where the block in thymocyte development is overcome and the transition between the CD4(-)CD8(-) and CD4(+)CD8(+) stages is fully restored. Determination of the expression levels of Fyn, ZAP-70 and Syk in thymocytes form the Lck(-/-) thymoma revealed high levels of ZAP-70 overexpression and recovery of a specific subset of phosphoproteins as compared to Lck(-/-) thymocytes. Hence ZAP-70 overexpression in thymocytes is associated with recovery from the developmental arrest caused by the absence of Lck, suggesting a role for ZAP-70 downstream of Lck in the maturation of CD4(+)CD8(+) thymocytes.     

Timing of deletion of autoreactive V{beta}6+ cells and down-modulation of either CD4 or CD8 on phenotypically distinct CD4+8+ subsets of thymocytes expressing intermediate or high levels of T cell receptor

In this paper we describe a differentiation sequence amongst adult murine thymocytes which goes from CD4+8+3lo(low) to CD4+8+3int(intermediate) to CD4+8+3hi(high) and then to mature single positive CD3hi thymocytes. Phenotypic characterization of CD4+8+3int/hi cells for a number of other surface markers is consistent with them being in transition from CD4+8+3lo phenotype to mature phenotype. The same observation was made for sensitivity towards ionomycin-mediated apoptosis. In the thymus of Mls-1a mice, where autoreactive TCR-V beta 6+ cells are negatively selected, deletion of TCR-V beta 6+ cells was first detected in the CD4+8+3int subset, and was complete by the CD4+8+3hi stage, suggesting that up-regulation of the TCR/CD3 complex is required for deletion of Mls-1a autoreactive thymocytes. No sign of apoptosis was detected among any fresh thymocyte subsets suggesting that apoptotic cells are rapidly cleared from the thymus. The CD4+8+3int/CD4+8+3hi cells are therefore populations in transit from the typical cortical thymocytes to the mature T-cells.     

Organotin-induced thymus atrophy concerns the OX-44+ immature thymocytes. Relation to the interaction between early thymocytes and thymic epithelial cells?

After single oral application of the organotin compound di-n-butyltindichloride (DBTC) to rats, a reversible dose-dependent thymus weight reduction is observed. This is maximal at day 4 and recovers to the control value approximately at day 9 after administration. In this study the changes in thymocyte subpopulations after a single oral dose of 15 mg DBTC/kg body weight were analysed by immunohistology. Thymus glands of exposed rats were collected at day 1,2,3,4,5,7 and 9 after DBTC dosing and frozen sections were screened for various thymocyte differentiation antigens. Staining by mAb HIS-44 that labels a subset of cortical thymocytes showed that the thymus atrophy was restricted to the cortex. Here a time-dependent decrease of labelling by CD2 (OX-34), CD8 (OX-8), CD4 (ER-2), and CD5 (OX-19) was observed. In contrast, the number of cortical OX-44+ cells increased from day 2 to day 5. This increase can reflect an increase of CD4-CD8- double-negative thymocytes or of macrophages. However, most of these OX-44+ cells were negative for acid phosphatase, which is present in most macrophages. We concluded that these OX-44+ cells were mainly CD4-CD8- thymocytes and that the thymocyte subpopulation of this phenotype, i.e. CD4-CD8-OX-44+, may be the target cell for DBTC. It is discussed whether DBTC might disturb the interaction of early thymocytes and thymic epithelium, probably by an interaction with the CD2 antigen.     

The Src-like adaptor protein downregulates the T cell receptor on CD4+CD8+ thymocytes and regulates positive selection

In this report, we show that the Src-like adaptor protein (SLAP) plays an important role in thymocyte development. SLAP expression is developmentally regulated; it is low in CD4-CD8- thymocytes, it peaks in the CD4+CD8+ subset, and it decreases to low levels in more mature cells. Disruption of the SLAP gene leads to a marked upregulation of TCR and CD5 expression at the CD4+CD8+ stage. The absence of SLAP was also developmentally significant because it enhanced positive selection in mice expressing the DO11.10 transgenic T cell receptor. Moreover, SLAP deletion at least partially rescued the development of ZAP-70-deficient thymocytes. These results demonstrate that SLAP participates in a novel mechanism of TCR downregulation at the CD4+CD8+ stage and regulates positive selection.