Involvement of the interleukin 2 pathway in the rearrangement and expression of both [alfa/beta] and [gamma/delta] T cell receptor genes in human T cell precursors (original) (raw)
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α β and γ δ T cells can share a late common precursor
Current Biology, 1995
The subdivision of T cells into o13 and y8 subtypes is conserved throughout vertebrate development. The respective ap and y8 T-cell receptors (TCRs) are encoded by somatically rearranged genes. There has been broad speculation as to whether an individual thymocyte can become either a y 8 T cell or an ao1 T cell as a result of stochastic gene rearrangement processes, or whether the two types of T cell are derived from separate lineages. Many of the experimental findings are apparently conflicting, however, and the issuea basic one in immunology and development -remains unresolved. Results: To address this issue, we have used the recently developed polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique, which allows us to examine quantitatively the status of TCR y and 8 genes in postnatal ap T cells and their progenitors. Interestingly, such cells are depleted of productively rearranged 8 and y genes, which can encode 8 and y TCR polypeptide chains. However, in mice that can rearrange TCR8 gene segments, but in which the TCR8 gene is non-functional in other respects, no such depletion of productive rearrangements is seen. Conclusion: The quantitative data that we have obtained fulfill the predictions of the stochastic hypothesis: that is, a progenitor T cell first attempts to become a /y 8 T cell and, if unsuccessful, then attempts to become an t13 T cell. Thus, oat3 and y8 T cells can derive from a common precursor thymocyte. In the simplest case, therefore, lineage-determining factors are the successful rearrangement of both y and 8 genes before TCRax gene rearrangements occur, which lead to deletion of the TCR8 locus and thereby preclude further y 8 T-cell differentiation. In contrast, successful rearrangement of the TCR3 locus remains compatible with cells becoming either y8 or ad3 T cells.
European Journal of Immunology, 1996
The earliest T precursor population in the adult mouse thymus, considered to have the surface phenotype CD4lo8−3−44+25−Thy‐1lo c‐kit+ (termed the low CD4 precursor), has been shown to have the capacity to produce B cells and dendritic cells, as well as T cells, and to have the T cell antigen receptor (TCR) Cβ gene region in germ‐line configuration. Because of evidence that this precursor population may have low levels of CD8 as well as CD4 on the cell surface, it was isolated, stained for surface CD4 and CD8 and assayed for the expression of messenger RNA (mRNA) for CD4 and CD8 by the reverse transcriptase polymerase chain reaction (RT‐PCR). The low CD4 precursors gave definite, moderate levels of staining for both CD8 and CD4, in contrast to downstream precursors which showed only marginal staining and so could be considered as genuine CD4−8−3− triple negatives. The low CD4 precursor expressed a significant level of mRNA for CD4, indicating that the surface CD4 was produced by thes...
Blood, 1999
Recent studies have identified several populations of progenitor cells in the human thymus. The hematopoietic precursor activity of these populations has been determined. The most primitive human thymocytes express high levels of CD34 and lack CD1a. These cells acquire CD1a and differentiate into CD4(+)CD8(+) through CD3(-)CD4(+)CD8(-) and CD3(-)CD4(+) CD8alpha+beta- intermediate populations. The status of gene rearrangements in the various TCR loci, in particular of TCRdelta and TCRgamma, has not been analyzed in detail. In the present study we have determined the status of TCR gene rearrangements of early human postnatal thymocyte subpopulations by Southern blot analysis. Our results indicate that TCRdelta rearrangements initiate in CD34(+)CD1a- cells preceding those in the TCRgamma and TCRbeta loci that commence in CD34(+)CD1a+ cells. Furthermore, we have examined at which cellular stage TCRbeta selection occurs in humans. We analyzed expression of cytoplasmic TCRbeta and cell-su...
A Molecular Map of T Cell Development
Immunity, 1998
; Fehling and von Boehmer, 1997). Early * Section of Immunology stages of thymocyte development are clearly defined Yale University School of Medicine by several molecular events that include expression of Howard Hughes Medical Institute the RAG-1 and RAG-2 gene products (Lin and Desiderio, New Haven, Connecticut 06520-8011 1993; Ferguson et al., 1994; Hoffman et al., 1996), so-† Laboratory of Immunology matic rearrangement of the T cell receptor  chain locus, National Institute of Allergy and Infectious Diseases expression of the coreceptor molecules CD4 and CD8, National Institutes of Health and, finally, recombination of the TCR ␣ locus (Dudley Bethesda, Maryland 20892 et al., 1994; Petrie et al., 1995). Productive rearrange-‡ Department of Molecular Biotechnology ment of both TCR chains allows surface expression of University of Washington School of Medicine the TCR and ensures life up to this point. Further devel-Seattle, Washington 98195-7650 opment, however, relies in part (Tourigny et al., 1997) upon signals generated by TCR interaction with MHC: self peptide complexes (von Boehmer, 1994). Most thy-Summary mocytes fail this rigorous criterion for selection and die of neglect (Janeway, 1994). Using a sensitive molecular marker for positive selec-While the early events in development can be clearly tion, the appearance of a particular functional TCR ␣ defined by molecular events, later stages of developchain sequence in cells from mice bearing a transgenic ment are typically defined by variations in the expression levels of cell surface proteins such as CD4, CD8, and  chain, we address several aspects of intrathymic T the TCR. These molecules allow for several different cell development. First, by examining specific TCR intrathymic subpopulations to be distinguished. The deprior to and after maturation, we demonstrate how a velopmental status of such populations has been, in restricted TCR repertoire is positively selected from most cases, assessed by other means including intraa highly diverse immature TCR repertoire. Second, thymic injections of genetically marked cells and reconsince this molecular marker is enriched in cells prostitution of thymocytes in irradiated mice. gressing toward the CD4 lineage and depleted in cells In this report, we identify a sensitive molecular marker progressing toward the CD8 lineage, a map of the for positive selection. This marker, which is the appeardevelopmental pathway of ␣ thymocytes can be inance or loss of a specific TCR ␣ chain sequence, is used ferred. Third, the first cells that show clear signs of to address three issues concerning the development of positive intrathymic selection are identified. TCR␣ bearing thymocytes. First, we demonstrate that the restriction of TCR usage that was apparent in the mature repertoire in "single-peptide" mice bearing a Peptide-Specific Responses of Nonimmunized TCR  Chain Transgenic Thymocytes § To whom correspondence should be addressed (e-mail: derek.
Characterization of T cell receptor (TCR) gene rearrangement during adult murine T cell development
The Journal of Immunology
Development of the ␣ and ␥␦ T cell lineages is dependent upon the rearrangement and expression of the TCR␣ and  or ␥ and ␦ genes, respectively. Although the timing and sequence of rearrangements of the TCR␣ and TCR loci in adult murine thymic precursors has been characterized, no similar information is available for the TCR␥ and TCR␦ loci. In this report, we show that approximately half of the total TCR␦ alleles initiate rearrangements at the CD44 high CD25 ؉ stage, whereas the TCR locus is mainly in germline configuration. In the subsequent CD44 low CD25 ؉ stage, most TCR␦ alleles are fully recombined, whereas TCR rearrangements are only complete on 10 -30% of alleles. These results indicate that rearrangement at the TCR␦ locus can precede that of TCR locus recombination by one developmental stage. In addition, we find a bias toward productive rearrangements of both TCR␦ and TCR␥ genes among CD44 high CD25 ؉ thymocytes, suggesting that functional ␥␦ TCR complexes can be formed before the rearrangement of TCR. These data support a model of lineage commitment in which sequential TCR gene rearrangements may influence ␣/␥␦ lineage decisions. Further, because TCR gene rearrangements are generally limited to T lineage cells, these analyses provide molecular evidence that irreversible commitment to the T lineage can occur as early as the CD44 high CD25 ؉ stage of development.
2000
We have recently identified a dominant wave of CD4-CD8-(double-negative [DN]) thymocytes in early murine fetal development that express low affinity Fc 3, receptors (FcyRII/III) and contain precursors for Tic~/3 lineage T cells. Here we show that Fc'yRII/III is expressed in very immature CD4 l~ single-positive (SP) thymocytes and that Fc3,RII/III expression is downregulated within the DN subpopulation and before the CD3-CD8 l~ SP stage in T call receptor (TCR)-a/3 lineage-committed thymocytes. DN Fc"t'RII/III + thymocytes also contain a small fraction of TCR-3'/t5 lineage cells in addition to TCR-c~/3 progenitors. Fetal day 15.5 DN TCR-a/B lineage progenitors can be subdivided into three major subpopulations as characterized by cell surface expression of Fc'yRII/III vs. CD2 (Fc'yRII/III+CD2 -, Fc3'RII/ III + CD2 +, Fc3,RII/III-CD2 +). Phenotypic analysis during fetal development as well as adoptive transfer of isolated fetal thymocyte subpopulations derived from C57B1/6 (Ly5.1) mice into normal, nonirradiated Ly5.2 congenic recipient mice identifies one early differentiation sequence (FcyRII/III + CD2-"-* Fc'yRII/III + CD2 + "-~ Fc3,RII/III-CD2 +) that precedes the entry of DN thymocytes into the CD4+CD8 + double-positive (DP) TCR l~ stage. Unseparated day 15.5 fetal thymocytes develop into DP thymocytes within 2.5 d and remain at the DP stage for >48 h before being selected into either CD4 + or CD8 + SP thymocytes. In contrast, Fc~/RII/ III+CD2 -DN thymocytes follow this same developmental pathway but are delayed by "~24 h before entering the DP compartment, while Fc'yRII/III-CD2 § display accelerated development by *24 h compared with total day 15.5 thymocytes. Fc'yRII/III-CD2 + are also more developmentally advanced than Fc3,RII/III+CD2 -fetal thymocytes with respect to their TCR ~/chain V(D)J rearrangement. At day 15.5 in gestation, ~ chain V(D)J rearrangement is mostly, if not entirely, restricted to the Fc3,RII/III-CD2 + subset of DN fetal thymocytes. Consistent with this analysis in fetal thymocytes, >90% of aduh thymocytes derived from mice carrying a disrupting mutation at the recombination-activating gene 2 locus (RAG-2-/-) on both alleles are developmentally arrested at the DN CD2-stage. In addition, there is a fivefold increase in the relative percentage of thymocytes expressing Fc3,RII/III in TCR and immunoglobulin gene rearrangementincompetent homozygous RAG-2-/-mice (15% Fc3,RII/III § versus rearrangement-competent heterozygous RAG-2 +/-mice (<3% FcyRII/III+). Thus, Fc3'RII/III expression defines an early DN stage preceding Vo(D~)Ja rearrangement, which in turn is followed by surface expression of CD2. Loss of Fc3,RII/III and acquisition of CD2 expression characterize a late DN stage immediately before the conversion into DP thymocytes.