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T Lindsten

Publications and source records attributed to T Lindsten.

At least 19 recordsLinked to original sources
PubMed1992

Regulation of translation initiation factor gene expression during human T cell activation.

Activation of quiescent T cells leads to a dramatic increase in the rate of protein synthesis. It is believed that this pronounced increase of protein synthesis is regulated primarily at the level of translational initiation. Although considerable evidence demonstrates that translational initiation can be regulated at the post-translational level by the phosphorylation/dephosphorylation of translation initiation factors (eIFs) such as eIF-4E and eIF-2 alpha, additional mechanisms of eIF gene expression may also play a role in the regulation of translation in quiescent cells and/or during their subsequent induction to enter the cell cycle. To address this issue, gene expression of eIF-2 alpha, -4E, and -4A was studied in quiescent human peripheral blood T cells following stimulation through the T cell receptor-CD3 complex. Quiescent T cells expressed low levels of eIF-2 alpha, -4E, and -4A mRNAs and proteins as compared to proliferating T cells. Activation of resting T cells resulted in a rapid increase (20-50-fold) in the levels of these three mRNAs. This increase did not require new protein synthesis. Furthermore, transcription rates of these three eIF genes showed only minor increase over the induction period as measured by nuclear run-on assays. Despite the rapid increase in initiation factor mRNA levels, increases in eIF protein levels lagged significantly behind. Western blot analysis also showed that the protein levels of the three eIFs were differentially increased. eIF-4A protein levels increased in proportion to the observed increase in cellular protein synthetic activity while the increases in eIF-4E and eIF-2 alpha proteins were proportionately less. The low levels of eIF proteins in quiescent T cells appear to correlate with low protein synthesis rate in such cells. The induction of eIF proteins by post-transcriptional/translational mechanisms appears to contribute to the pronounced stimulation of protein synthesis that occurs during T cell activation.

Animals↗
PubMed1992

AU RNA-binding factors differ in their binding specificities and affinities.

AUUUA multimers present in the 3'-untranslated region of mature lymphokine and cytokine transcripts have been implicated in the regulation of mRNA stability and translational efficiency. We have identified RNA-binding factors, termed AU-A, AU-B, and AU-C, that interact with AUUUA multimers. AU-A is an abundant, constitutively expressed 34-kDa factor that localizes primarily to the nucleus. AU-A binds to AUUUA multimers with low relative affinity and also binds to other U-rich sequences, including a poly(U) sequence. AU-B and AU-C are 30- and 43-kDa cytoplasmic factors that are induced following T cell receptor-mediated stimulation of purified human T cells and bind to AUUUA multimers with high affinity. Protease cleavage of AU-A, AU-B, and AU-C RNA-protein complexes indicate that AU-B and AU-C are structurally related to each other but distinct from AU-A. AU-B and AU-C require three or more tandem AUUUA repeats for efficient binding, and binding by these factors poorly tolerates mutations in the AUUUA recognition sequence. The precise binding specificity, high affinity, pattern of induction, and cytoplasmic localization all suggest that the structurally related AU-B and AU-C RNA-binding factors could be cytoplasmic regulators of lymphokine mRNA metabolism.

Antigens, CD↗
PubMed1992

Interleukin-4 differentially regulates tumor necrosis factor-alpha gene expression by human T lymphocytes and monocytes.

Tumor necrosis factor-alpha (TNF-alpha), a product of both mononuclear phagocytes and T lymphocytes, is an important proximal mediator of a number of acute and chronic inflammatory disease states. In this investigation we examine the regulatory effects of the lymphocyte product interleukin-4 (IL-4) on the gene expression of TNF-alpha from stimulated human peripheral blood monocytes (PBM) and T lymphocytes. We demonstrated the dose-dependent suppression of TNF-alpha mRNA and protein synthesis from lipopolysaccharide-treated PBM by IL-4. The suppressive effects of IL-4 appear to be dependent upon de novo protein synthesis, as cycloheximide abrogated the IL-4-induced reduction in TNF-alpha mRNA levels from PBM. In contrast to the suppressive effects of IL-4 on PBM-derived cytokine expression, IL-4 did not alter TNF-alpha mRNA expression from alpha-Cd3 or PMA + alpha-CD-28-treated T lymphocytes. Moreover, IL-2 mRNA expression from similarly treated T lymphocytes was unaltered by IL-4. Our findings demonstrate that disparity exists in the regulation of TNF-alpha gene expression from different immune cell populations which may have important implications in the evolution of acute and chronic inflammatory responses.

Base Sequence↗
PubMed1991

An inducible cytoplasmic factor (AU-B) binds selectively to AUUUA multimers in the 3' untranslated region of lymphokine mRNA.

Considerable evidence suggests that the metabolism of lymphokine mRNAs can be selectively regulated within the cytoplasm. However, little is known about the mechanism(s) that cells use to discriminate lymphokine mRNAs from other mRNAs within the cytoplasm. In this study we report a sequence-specific cytoplasmic factor (AU-B) that binds specifically to AUUUA multimers present in the 3' untranslated region of lymphokine mRNAs. AU-B does not bind to monomeric AUUUA motifs nor to other AU-rich sequences present in the 3' untranslated region of c-myc mRNA. AU-B RNA-binding activity is not present in quiescent T cells but is rapidly induced by stimulation of the T-cell receptor/CD3 complex. Induction of AU-B RNA-binding activity requires new RNA and protein synthesis. Stabilization of lymphokine mRNA induced by costimulation with phorbol myristate acetate correlates inversely with binding by AU-B. Together, these data suggest that AU-B is a cytoplasmic regulator of lymphokine mRNA metabolism.

Base Sequence↗
PubMed1990

Sequence-specific binding of human Ets-1 to the T cell receptor alpha gene enhancer.

Expression of the human T cell receptor (TCR) alpha gene is regulated by a T cell-specific transcriptional enhancer that is located 4.5 kilobases (kb) 3' to the C alpha gene segment. The core enhancer contains two nuclear protein binding sites, T alpha 1 and T alpha 2, which are essential for full enhancer activity. T alpha 1 contains a consensus cyclic adenosine monophosphate (cAMP) response element (CRE) and binds a set of ubiquitously expressed CRE binding proteins. In contrast, the transcription factors that interact with the T alpha 2 site have not been defined. In this report, a lambda gt11 expression protocol was used to isolate a complementary DNA (cDNA) that programs the expression of a T alpha 2 binding protein. DNA sequence analysis demonstrated that this clone encodes the human ets-1 proto-oncogene. Lysogen extracts produced with this cDNA clone contained a beta-galactosidase-Ets-1 fusion protein that bound specifically to a synthetic T alpha 2 oligonucleotide. The Ets-1 binding site was localized to a 17-base pair (bp) region from the 3' end of T alpha 2. Mutation of five nucleotides within this sequence abolished both Ets-1 binding and the activity of the TCR alpha enhancer in T cells. These results demonstrate that Ets-1 binds in a sequence-specific fashion to the human TCR alpha enhancer and suggest that this developmentally regulated proto-oncogene functions in regulating TCR alpha gene expression.

Base Sequence↗
PubMed1990

CD28 ligation in T-cell activation: evidence for two signal transduction pathways.

The CD28 homodimer is thought to function as a signal transducing receptor during activation of T cells. Evidence is presented that the degree of aggregation of CD28 on the cell surface regulates two distinct CD28-associated signals. Binding of bivalent CD28 monoclonal antibody (MoAb) 9.3 upregulates lymphokine production by messenger RNA (mRNA) stabilization, without direct initiation of lymphokine mRNA transcription. This signal was not dependent on inositol phospholipid production or activation of a protein tyrosine kinase (PTK). In contrast, further crosslinking of CD28 on the cell surface rapidly induced formation of large amounts of inositol trisphosphate (InsP3) and increased cytoplasmic calcium concentration [( Ca2+]i), but did not stimulate PTK. CD28 crosslinking directly activated a subset of resting T cells, since CD25 (interleukin [IL]-2 receptor alpha chain) mRNA was rapidly induced in purified T cells, and proliferation, even without addition of exogenous IL-2, was sometimes observed. CD25 expression was detected on the cell surface of approximately 20% of CD4+ T cells. The degree of CD28 aggregation required for activation was investigated by preparing soluble 9.3 x 9.3 conjugates ranging in size from approximately 300 Kd to greater than 1,000 Kd, and comparing their function in T-cell proliferation assays with phorbol-12-myristate-13-acetate (PMA), anti-CD3, or IL-2. There was a correlation between conjugate size and proliferation with IL-2, whereas costimulation with PMA or CD3 was optimized at a lower degree of CD28 aggregation. The inositol phospholipid (InsP) generation and increase in [Ca2+]i after CD28 receptor aggregation appeared to proceed through a pathway different from the CD3/T-cell receptor (TCR) pathway since it was enhanced by pretreatment with PMA, while the InsP and [Ca2+]i signal from crosslinking CD3 was suppressed by PMA. Furthermore, the proliferation response to CD28 aggregation was resistant to inhibition by CD3 modulation. Thus, CD28 aggregation appears to trigger a phospholipase C activation pathway that differs from the CD3/TCR-linked pathway.

Animals↗
PubMed1990

Reciprocal expression of human ETS1 and ETS2 genes during T-cell activation: regulatory role for the protooncogene ETS1.

The expression of the protooncogenes ETS1 and ETS2 has been studied in purified human T cells activated either by cross-linking of the T-cell receptor-CD3 complex on their cell surface or by direct stimulation with phorbol esters and ionomycin. Our results show that resting T cells express high levels of ETS1 mRNA and protein, while expression of ETS2 is undetectable. Upon T-cell activation, ETS2 mRNA and proteins are induced, while ETS1 gene expression decreases to very low levels. Late after stimulation, ETS1 mRNA is reinduced and maintained at a high level, while ETS2 gene expression decreases to undetectable levels. Therefore, it appears that in human T cells, ETS2 gene products are associated with cellular activation and proliferation, while ETS1 gene products are preferentially expressed in a quiescent state.

Calcium↗
PubMed1989

Evidence for the involvement of three distinct signals in the induction of IL-2 gene expression in human T lymphocytes.

The regulation of IL-2 gene expression during T cell activation and proliferation has been investigated in primary cultures of purified human peripheral blood T cells. Prior results indicated that stimulation of T cells by anti-CD28 mAb plus PMA could induce IL-2 expression and T cell proliferation that was entirely resistant to cyclosporine. The present studies examined whether CD28 augments IL-2 expression by a unique pathway or merely acts at a point common to CD3-induced proliferation but distal to the effects of cyclosporine. The induction of maximal IL-2 gene expression required three signals provided by phorbol ester, calcium ionophore, and anti-CD28 mAb. Stimulation of cells by optimal amounts of calcium ionophore and PMA induced IL-2 mRNA that was completely suppressed by cyclosporine. The addition of anti-CD28 to T cells stimulated with PMA plus calcium ionophore induced a 5- to 100-fold increase in IL-2 gene expression and secretion that was resistant to cyclosporine. The CD28 signal was able to increase steady state IL-2 mRNA levels even in cells treated with maximally tolerated amounts of calcium ionophore and PMA. The three-signal requirement did not reflect differential regulation of lymphokine gene expression between the CD4 and CD8 T cell subsets or differences in the kinetics of IL-2 mRNA expression. The signal provided by CD28 is distinct from that of CD3 because although anti-CD28 plus PMA-induced proliferation is resistant to cyclosporine, anti-CD3 or anti-CD3 plus PMA-induced IL-2 expression is sensitive. Thus, these studies show that three biochemically distinct signals are required for maximal IL-2 gene expression. Furthermore, these studies suggest that lymphokine production in T cells is not controlled by an "on/off" switch, but rather, that CD28 regulates a distinct intracellular pathway which modulates the level of IL-2 production on a per cell basis. The observation that CD28 stimulation results in IL-2 concentrations that exceed 1000 U/m1 in tissue culture supernatants suggests that a role in vivo for CD28 might be to amplify immune responses initiated by the CD3/T cell receptor complex. Finally, the observation that CD28 interacts with the signals provided by PMA and calcium ionophore shows that the function of CD28 is not merely to act as a scaffold to stabilize or enhance signalling through the CD3/TCR complex.

Antibodies, Monoclonal↗
PubMed1989

Two distinct mechanisms of interleukin-2 gene expression in human T lymphocytes.

Interleukin-2 (IL-2) gene regulation was investigated in primary cultures of highly purified human peripheral blood CD28+ T cells. Two discrete mechanisms for induction of T-cell proliferation could be distinguished by examining cell cycle progression and the expression of the IL-2 gene. Stimulation of cells by CD3 MoAb induced only transiently expressed, small amounts of IL-2 mRNA that was completely suppressed by cyclosporine. Costimulation of T cells with CD3 MoAb and either CD28 MoAb or PMA, but not calcium ionophore, induced a 50-100-fold increased in IL-2 gene expression and secretion. High levels of IL-2 gene expression could also be achieved by stimulation with calcium ionophore and PMA or CD28 MoAb and PMA, but not by CD28 MoAb plus calcium ionophore. IL-2 gene expression and T-cell proliferation induced by CD3 MoAb plus PMA or calcium ionophore plus PMA were completely suppressible by cyclosporine. In contrast, IL-2 gene expression and T-cell proliferation induced by CD28 MoAb plus PMA were unaffected by cyclosporine. The CD28 signal was dependent on new protein synthesis. Nuclear run-on transcription assays showed that anti-CD28 did not affect lymphokine transcription. A major effect of CD28 stimulation on mRNA stability was shown by studies using actinomycin D; CD28 stimulation substantially increased the half-life of IL-2 and TNF-alpha mRNA. The effects of anti-CD28 stimulation were specific for growth factors, and thus differ from previously described effects of cycloheximide on mRNA stability. These studies suggest the existence of two biochemical pathways for the induction of IL-2 production, one that occurs at the transcriptional level and is mediated by intracellular calcium release and protein kinase C and is cyclosporine-sensitive, and one that acts post-transcriptionally, is mediated by CD28 stimulation, and is cyclosporine-resistant.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1989

The 4F2 heavy chain gene: a molecular model of inducible gene expression in human T cells.

We have utilized the human 4F2 heavy chain (4F2HC) gene as a model system in studies designed to elucidate the molecular events involved in regulating inducible gene expression during normal human T-cell activation. In previous studies we have shown that steady state levels of 4F2HC mRNA are induced 50-60-fold within 6 h of T-cell activation by phytohemagluttinin (PHA) and that the induction of 4F2HC gene expression involves both the protein kinase C and calcium-mediated activation pathways. Despite the fact that the 4F2HC gene is highly regulated in T cells, the 5' upstream region of the 4F2HC gene contains a housekeeping promoter which is G + C rich, lacks TATA or CCAAT sequences, and contains four potential binding sites for the ubiquitous Sp 1 transcription factor. The major regulatory elements of the 4F2HC gene do not reside within this 5' upstream region but instead, map to the exon 1-intron 1 region of the gene. The low levels of mature 4F2HC mRNA in resting T cells result from a block to transcription elongation within the exon 1-intron 1 region of the gene rather than promoter inactivity. Phorbol ester stimulation of resting T cells induces 4F2HC expression by removing this block to transcription elongation. We now report that in addition to its ability to serve as a transcriptional attenuator, the 4F2HC first intron contains a powerful enhancer element which is active in a wide variety of cell types including malignant human T cells. Full enhancer activity is displayed by a 186 bp fragment of the first intron which contains binding sites for two novel nuclear proteins (NF-4FA and NF-4FB) which flank a consensus binding site for the AP-1 transcription factor. A cDNA encoding the NF-4FB enhancer binding protein has been cloned by screening a lambda gt11 cDNA library with a rabiolabelled oligonucleotide corresponding to the NF-4FB recognition sequence.

Antigens, Surface↗
PubMed1989

CD28 activation pathway regulates the production of multiple T-cell-derived lymphokines/cytokines.

CD28 is a 44-kDa glycoprotein expressed as a homodimer on the surface of a major subset of human T cells. Previous studies have demonstrated that the binding of monoclonal antibodies to the CD28 surface antigen can augment the proliferation of purified human T cells stimulated with suboptimal doses of mitogens or anti-T-cell receptor/CD3 complex antibodies. In this report, we show that CD28 stimulation augments T-cell immune responses by specifically inducing a 5- to 50-fold enhancement in the expression and secretion of interleukin 2, tumor necrosis factor type alpha, lymphotoxin, interferon gamma, and granulocyte-macrophage colony-stimulating factor in normal human T cells stimulated to proliferate by crosslinking of the T-cell receptor/CD3 complex. This CD28-mediated induction of lymphokine/cytokine gene expression occurred even in T cells stimulated with optimal concentrations of mitogens or anti-T-cell receptor/CD3 antibodies, although under these conditions CD28 activation failed to enhance the proliferative response. The activation pathway induced by stimulation of CD28 is distinct from other biochemical pathways that induce lymphokines/cytokines because CD28 stimulation can induce lymphokine/cytokine gene expression in the presence of the immunosuppressant cyclosporine. Together these data suggest that the CD28 cell surface molecule is part of a distinct activation pathway that specifically modulates the expression of multiple lymphokine/cytokine genes.

Antibodies, Monoclonal↗
PubMed1988

Transcription of T cell antigen receptor genes is induced by protein kinase C activation.

The regulation of expression of the TCR-alpha and -beta genes was studied in the human T cell tumor line Jurkat. Treatment of the cells with PMA was shown to decrease the surface expression of the TCR-alpha/beta/CD3 complex. Subsequent to PMA-induced modulation of the TCR/CD3 complex, increases in the mRNA levels of both the TCR-alpha and -beta genes were observed reaching a maximum 12 h after stimulation. Other T cell activators were also examined for their ability to increase TCR-alpha and -beta mRNA expression. Only agents that activate protein kinase C were shown to induce expression of the TCR-alpha and -beta genes. The observed increases in TCR-alpha and -beta gene mRNA levels were not the result of a uniquely derived Jurkat subline. Similar inductions of TCR-alpha and -beta mRNA levels were observed in an independently maintained Jurkat cell line. In both cell lines, elevations of TCR gene expression was accompanied by a decline in the expression of the c-myc proto-oncogene. PMA induction of TCR-alpha and -beta mRNA was shown to occur in the presence of the protein synthesis inhibitor cycloheximide. The 1.6-kb TCR-alpha and the 1.0-kb D beta J beta C beta TCR-beta gene transcripts were fully induced in the presence of cycloheximide, whereas the 1.3-kb V beta D beta J beta C beta transcript was only partly induced in the presence of cycloheximide. Run-on transcription assays demonstrated that the increase in TCR-alpha and -beta mRNA levels could be entirely accounted for by increases in the transcription rate of both genes after PMA induction. Thus, in summary, protein kinase C stimulation leads to TCR-alpha/beta modulation in Jurkat cells and an increase in steady state TCR-alpha and -beta mRNA levels as a result of transcriptional activation of both genes.

Cell Line↗
PubMed1988

Multiple mechanisms regulate c-myc gene expression during normal T cell activation.

Quiescent normal human T cells express low levels of steady-state c-myc mRNA as a result of low constitutive promoter utilization, a block to transcriptional elongation within the gene, and rapid degradation of c-myc mRNA in the cytoplasm. Following the activation of the T cell receptor (TCR)/CD3 complex, quiescent T cells are induced to express c-myc mRNA. Two intracellular pathways, one involving protein kinase C activation and the other mediated by increased intracellular calcium concentration, are activated by TCR/CD3 receptor stimulation. These two pathways, which can be activated by phorbol myristate acetate (PMA) and ionomycin respectively, appear to play complementary roles in the transcriptional induction of c-myc gene expression by the antigen receptor complex. Ionomycin treatment of quiescent cells leads to enhanced c-myc expression primarily as a result of increased transcriptional initiation. In contrast, PMA contributes to c-myc expression, at least in part, by decreasing the block to transcriptional elongation present within the gene. Both the PMA- and ionomycin-mediated induction of c-myc expression can be independently enhanced by stabilization of c-myc mRNA in the cytoplasm. These observations demonstrate that multiple mechanisms co-operate to regulate c-myc gene expression during normal T cell activation.

Anti-Bacterial Agents↗
PubMed1988

Isolation and structural characterization of the human 4F2 heavy-chain gene, an inducible gene involved in T-lymphocyte activation.

The human 4F2 cell surface antigen is a 120-kilodalton (kDa) disulfide-linked heterodimer which is composed of an 80- to 90-kDa glycosylated heavy chain (4F2HC) and a 35- to 40-kDa nonglycosylated light chain (4F2LC). 4F2 belongs to a family of inducible cell surface molecules which are involved in T-lymphocyte activation and growth. To better understand the molecular mechanism(s) that controls 4F2HC gene expression in both resting and activated T cells, a 4F2HC human genomic clone was isolated and structurally characterized. The 4F2HC gene spans 8 kilobases of chromosome 11 and is composed of nine exons. The 5' upstream region of the gene displays several properties which are characteristic of housekeeping genes. It is G+C rich and hypomethylated in peripheral blood lymphocyte DNA and contains multiple binding sites for the Sp1 transcription factor while lacking TATA or CCAAT sequences. This region of the gene also displays sequence homologies with several other inducible T-cell genes, including the interleukin-2, interleukin-2 receptor alpha chain, dihydrofolate reductase, thymidine kinase, and transferrin receptor genes. A 255-base-pair fragment of the 4F2HC gene which contains 154 base pairs of the 5' flanking sequence was able to efficiently promote expression of the bacterial chloramphenicol acetyltransferase gene in human Jurkat T cells, indicating that it contains promoter or enhancer (or both) sequences. Analyses of chromatin structure in resting and lectin-activated T cells revealed the presence of stable DNase I-hypersensitive sites within both the 5' flanking and intron 1 regions of the 4F2HC gene. Although the 4F2HC gene displayed many of the structural features characteristic of a constitutively expressed gene, lectin-mediated activation of resting peripheral blood T lymphocytes resulted in a dramatic increase in steady-state levels of 4F2HC mRNA.

Amino Acid Sequence↗
PubMed1988

Regulation of 4F2 heavy-chain gene expression during normal human T-cell activation can be mediated by multiple distinct molecular mechanisms.

The 4F2 molecule belongs to the set of cell surface antigens which is induced following lectin- or antigen-mediated T-cell activation. The increase in 4F2 cell surface expression following lectin-mediated stimulation has been shown to be accompanied by a parallel increase in the steady-state levels of 4F2 heavy-chain (4F2HC) mRNA. The studies described in this report were designed to further elucidate the molecular mechanisms responsible for induction of 4F2HC gene expression following activation of normal resting human peripheral blood T cells. The low levels of mature 4F2HC mRNA in resting T cells were shown to be the result of a block to transcription elongation within the exon 1-intron 1 region of the 4F2HC gene rather than promoter inactivity. Phorbol myristate acetate stimulation of resting T cells resulted in a 20-fold increase in steady-state 4F2HC mRNA levels which was mediated by removal of this block to transcription elongation. The phorbol myristate acetate-induced increase in 4F2HC gene expression is distinct from previously described AP-1-mediated, phorbol ester-induced gene expression in that it requires new protein synthesis. Treatment of resting T cells with ionomycin plus PMA resulted in a 60-fold increase in 4F2HC mRNA levels. This induction was mediated by both an increase in promoter utilization and removal of the block to transcription elongation. Finally, by increasing the half-life of 4F2HC mRNA, cycloheximide treatment of resting T cells induced an approximately five fold increase in the levels of 4F2HC gene expression, although the physiologic significance of this mechanism remains unclear. These results demonstrate that the level of 4F2HC gene expression in normal peripheral blood T cells can be regulated by at least three distinct molecular pathways: (i) changes in promoter utilization, (ii) modulation of a block to transcription elongation, and (iii) alteration in mRNA stability.

Antigens, Surface↗
PubMed1987

Rearrangement and expression of T cell receptor genes in cloned murine natural suppressor cell lines.

Naturally occurring suppressor cells of the in vitro mixed leukocyte culture reaction and of in vivo graft-vs.-host disease have been identified in the spleens of neonatal mice (1) and of adult mice recovering from total lymphoid irradiation (2), whole-body irradiation (3), and syngeneic marrow transplantation (4), or cyclophosphamide therapy (5). Using both positive and negative selection procedures, the suppressors were reported to be null lymphocytes that did not express mature macrophage surface markers, nor differentiate into mature macrophages in vitro, nor demonstrate natural killer (NK) activity (1). Subsequently, cloned lines of these natural suppressor (NS) cells were derived from either adult mice given total lymphoid irradiation (TLI) (2) or from neonates (6). The cloned NS cell lines expressed a surface phenotype (2, 6) similar to that reported previously for cloned NK cells (Thy-1(+), asialo-GM1(+), Ig(-), Lyt-1(-), Lyt-2(-), Ia(-), MAC-1(-)) (7-9). However, the NS cells did not show NK activity in the standard assay with YAC-1 target cells. The cloned NS lines suppressed the proliferation of responder cells and the generation of cytolytic cells in the mixed leukocyte reaction (MLR), and suppressed lethal graft-vs.-host disease in vivo (10, 11). In view of the unusual function and surface phenotype of the cells, the lineage of these cells remained unclear. To determine the lineage of the cloned NS cells, we searched for expression and rearrangement of the alpha and beta chain genes of the T cell antigen receptor, as well as that of the gamma chain gene. Studies of the phenotypically similar NK cell yielded conflicting results. Thus, cloned lines of murine NK cells were reported to have rearrangements of the beta chain genes, and to express mRNA for all three chains (12). In contrast, freshly purified rat or human large granular lymphocytes (LGL) were shown to express only the 1.0 kb mRNA species of the beta chain gene (13), indicative of D-J joining (14). Thus, some but not all cells with NK function express the T cell receptor and are members of the T cell lineage. The current report shows that the NS lines express full-length mRNA transcripts for the a and beta chain of the T cell receptor, as well as the gamma chain gene.

Animals↗
PubMed1987

Transient rearrangements of the T cell antigen receptor alpha locus in early thymocytes.

The dull Ly-1 double-negative (Ly-1dull, Lyt-2-, L3T4-) subpopulation appears to be the major precursor group of T lymphocytes in the thymus. In examining the status of the alpha, beta, and gamma chain genes for T cell receptors (TCR) in this population of cells and hybridomas made from them, we find that all of these loci appear to begin DNA rearrangements in a nearly simultaneous fashion. In the case of the gamma genes, these involve V gamma----J gamma C gamma gene rearrangements; with the beta chain genes, both D beta----J beta C beta rearrangement and V beta----D beta J beta C beta rearrangements are evident; and in the case of the alpha locus, assayed in part by pulsed-field gel electrophoresis, they take the form of a novel series of rearrangements occurring 80 kb or more 5' to the C alpha gene. These alpha locus rearrangements are well away from any of the J alpha gene segments found in cDNA clones to date and are deleted in most mature thymocytes and functional T cell lines. Therefore they appear to represent a distinct class of rearrangement that occurs before V alpha----J alpha joining. These distinctions between the character of the TCR gene rearrangements in these cells represent useful markers in further distinguishing different stages of T cell differentiation within this compartment of early T cells. In addition, the unexpected discovery of clonal rearrangements so far away from any of the expressed J alpha gene segments, and at a stage where there is little or no stable C alpha RNA present, has interesting implications for the hierarchy of TCR gene expression.

Animals↗