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Commitment of immature CD4+8+ thymocytes to the CD4 lineage requires CD3 signaling but does not require expression of clonotypic T cell receptor (TCR) chains.

Suzuki H, Shinkai Y, Granger LG, Alt FW, Love PE, Singer A - J. Exp. Med. (1997)

Bottom Line: As a consequence of positive selection in the thymus, immature CD4(+)8(+) double-positive, [DP] thymocytes selectively terminate synthesis of one coreceptor molecule and, as a result, differentiate into either CD4(+) or CD8(+) T cells.Previously, we reported that the intrathymic signals that induced commitment to the CD4 versus CD8 T cell lineages were markedly asymmetric.Notably, CD8 commitment appeared to require lineage-specific signals, whereas CD4 commitment appeared to occur in the absence of lineage-specific signals by default.

View Article: PubMed Central - PubMed

Affiliation: Experimental Immunology Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.

ABSTRACT
As a consequence of positive selection in the thymus, immature CD4(+)8(+) double-positive, [DP] thymocytes selectively terminate synthesis of one coreceptor molecule and, as a result, differentiate into either CD4(+) or CD8(+) T cells. The decision by individual DP thymocytes to terminate synthesis of one or the other coreceptor molecule is referred to as lineage commitment. Previously, we reported that the intrathymic signals that induced commitment to the CD4 versus CD8 T cell lineages were markedly asymmetric. Notably, CD8 commitment appeared to require lineage-specific signals, whereas CD4 commitment appeared to occur in the absence of lineage-specific signals by default. Consequently, it was unclear whether CD4 commitment, as revealed by selective termination of CD8 coreceptor synthesis, occurred in all DP thymocytes, or whether CD4 commitment occurred only in T cell receptor (TCR)-CD3-signaled DP thymocytes. Here, we report that selective termination of CD8 coreceptor synthesis does not occur in DP thymocytes spontaneously. Rather, CD4 commitment in DP thymocytes requires signals transduced by either CD3 or zeta chains, which can signal CD4 commitment even in the absence of clonotypic TCR chains.

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CD5 surface expression on thymocytes from experimental  mice. Thymocytes from the indicated experimental mice were stained  with anti-CD5 mAb (solid line) or an irrelevant antibody (shaded curve).  CD5 expression on normal B6 thymocytes stained at the same time as a  positive control is also shown for comparison (dotted line).
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Figure 2: CD5 surface expression on thymocytes from experimental mice. Thymocytes from the indicated experimental mice were stained with anti-CD5 mAb (solid line) or an irrelevant antibody (shaded curve). CD5 expression on normal B6 thymocytes stained at the same time as a positive control is also shown for comparison (dotted line).

Mentions: To assess directly the role of TCR–CD3 signals in inducing lineage commitment in DP thymocytes, we assessed DP thymocytes from experimentally induced RAG2° mice. RAG2° mice fail to express any clonotypic TCR chain because they are unable to recombine any TCR gene locus. As a result, RAG2° thymocytes are arrested at the CD4−8− (double-negative, DN) stage of development (23). However, RAG° thymocytes can be induced to differentiate further into DP cells by either sublethal γ-irradiation (26, 27) or by injection of anti-CD3ε mAb (28, 29) (Fig. 1, A and B, left column). Induced RAG° DP thymocytes did not further differentiate into phenotypically mature T cells as thymocytes from stimulated RAG° mice that appeared CD4+8− or CD4−8+ (Fig. 1, A and B, left columns) were predominantly precursor cells that spontaneously became CD4+8+ in overnight culture (data not shown; reference 30). Even though sublethal γ-irradiation and anti-CD3ε injection both induced generation of RAG° DP thymocytes (Fig. 1), the two modes of stimulation did not have identical effects. That is, sublethal γ-irradiation did not detectably stimulate CD3 signal transduction as revealed by the absence of CD5 upregulation, whereas injection of anti-CD3ε mAb did stimulate CD3 signal transduction and CD5 upregulation (Fig. 2). Assessment of CD4+8lo and CD4lo8+ sorted cells from γ-irradiated RAG2° mice by the coreceptor reexpression assay revealed that none had undergone lineage commitment (Fig. 1, A and B). In contrast, assessment of CD4+8lo sorted cells from anti-CD3ε– induced RAG2° mice revealed the presence of CD4-committed DP thymocytes that had selectively terminated CD8 coreceptor synthesis, as well as the presence of uncommitted DP thymocytes (Fig. 1 A). The CD4-committed thymocytes that were present in anti-CD3ε–induced RAG2° mice expressed the phenotype of newly committed thymocytes in that they were HSAhi, thymic shared antigen (TSA)-1hi (data not shown). In contrast, anti-CD3ε–induced RAG2° thymocytes had no CD8-committed cells among either CD4+8lo or CD4lo8+ sorted cell populations (Fig. 1, A and B). Thus, these results (a) confirm that DP thymocytes do not undergo lineage commitment in the absence of TCR–CD3 signals, and (b) demonstrate that CD3 signals stimulated by anti-CD3ε mAbs are sufficient to induce DP thymocytes to selectively terminate CD8 coreceptor synthesis and commit to the CD4 lineage, even in the absence of clonotypic TCR chains.


Commitment of immature CD4+8+ thymocytes to the CD4 lineage requires CD3 signaling but does not require expression of clonotypic T cell receptor (TCR) chains.

Suzuki H, Shinkai Y, Granger LG, Alt FW, Love PE, Singer A - J. Exp. Med. (1997)

CD5 surface expression on thymocytes from experimental  mice. Thymocytes from the indicated experimental mice were stained  with anti-CD5 mAb (solid line) or an irrelevant antibody (shaded curve).  CD5 expression on normal B6 thymocytes stained at the same time as a  positive control is also shown for comparison (dotted line).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2198962&req=5

Figure 2: CD5 surface expression on thymocytes from experimental mice. Thymocytes from the indicated experimental mice were stained with anti-CD5 mAb (solid line) or an irrelevant antibody (shaded curve). CD5 expression on normal B6 thymocytes stained at the same time as a positive control is also shown for comparison (dotted line).
Mentions: To assess directly the role of TCR–CD3 signals in inducing lineage commitment in DP thymocytes, we assessed DP thymocytes from experimentally induced RAG2° mice. RAG2° mice fail to express any clonotypic TCR chain because they are unable to recombine any TCR gene locus. As a result, RAG2° thymocytes are arrested at the CD4−8− (double-negative, DN) stage of development (23). However, RAG° thymocytes can be induced to differentiate further into DP cells by either sublethal γ-irradiation (26, 27) or by injection of anti-CD3ε mAb (28, 29) (Fig. 1, A and B, left column). Induced RAG° DP thymocytes did not further differentiate into phenotypically mature T cells as thymocytes from stimulated RAG° mice that appeared CD4+8− or CD4−8+ (Fig. 1, A and B, left columns) were predominantly precursor cells that spontaneously became CD4+8+ in overnight culture (data not shown; reference 30). Even though sublethal γ-irradiation and anti-CD3ε injection both induced generation of RAG° DP thymocytes (Fig. 1), the two modes of stimulation did not have identical effects. That is, sublethal γ-irradiation did not detectably stimulate CD3 signal transduction as revealed by the absence of CD5 upregulation, whereas injection of anti-CD3ε mAb did stimulate CD3 signal transduction and CD5 upregulation (Fig. 2). Assessment of CD4+8lo and CD4lo8+ sorted cells from γ-irradiated RAG2° mice by the coreceptor reexpression assay revealed that none had undergone lineage commitment (Fig. 1, A and B). In contrast, assessment of CD4+8lo sorted cells from anti-CD3ε– induced RAG2° mice revealed the presence of CD4-committed DP thymocytes that had selectively terminated CD8 coreceptor synthesis, as well as the presence of uncommitted DP thymocytes (Fig. 1 A). The CD4-committed thymocytes that were present in anti-CD3ε–induced RAG2° mice expressed the phenotype of newly committed thymocytes in that they were HSAhi, thymic shared antigen (TSA)-1hi (data not shown). In contrast, anti-CD3ε–induced RAG2° thymocytes had no CD8-committed cells among either CD4+8lo or CD4lo8+ sorted cell populations (Fig. 1, A and B). Thus, these results (a) confirm that DP thymocytes do not undergo lineage commitment in the absence of TCR–CD3 signals, and (b) demonstrate that CD3 signals stimulated by anti-CD3ε mAbs are sufficient to induce DP thymocytes to selectively terminate CD8 coreceptor synthesis and commit to the CD4 lineage, even in the absence of clonotypic TCR chains.

Bottom Line: As a consequence of positive selection in the thymus, immature CD4(+)8(+) double-positive, [DP] thymocytes selectively terminate synthesis of one coreceptor molecule and, as a result, differentiate into either CD4(+) or CD8(+) T cells.Previously, we reported that the intrathymic signals that induced commitment to the CD4 versus CD8 T cell lineages were markedly asymmetric.Notably, CD8 commitment appeared to require lineage-specific signals, whereas CD4 commitment appeared to occur in the absence of lineage-specific signals by default.

View Article: PubMed Central - PubMed

Affiliation: Experimental Immunology Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.

ABSTRACT
As a consequence of positive selection in the thymus, immature CD4(+)8(+) double-positive, [DP] thymocytes selectively terminate synthesis of one coreceptor molecule and, as a result, differentiate into either CD4(+) or CD8(+) T cells. The decision by individual DP thymocytes to terminate synthesis of one or the other coreceptor molecule is referred to as lineage commitment. Previously, we reported that the intrathymic signals that induced commitment to the CD4 versus CD8 T cell lineages were markedly asymmetric. Notably, CD8 commitment appeared to require lineage-specific signals, whereas CD4 commitment appeared to occur in the absence of lineage-specific signals by default. Consequently, it was unclear whether CD4 commitment, as revealed by selective termination of CD8 coreceptor synthesis, occurred in all DP thymocytes, or whether CD4 commitment occurred only in T cell receptor (TCR)-CD3-signaled DP thymocytes. Here, we report that selective termination of CD8 coreceptor synthesis does not occur in DP thymocytes spontaneously. Rather, CD4 commitment in DP thymocytes requires signals transduced by either CD3 or zeta chains, which can signal CD4 commitment even in the absence of clonotypic TCR chains.

Show MeSH
Related in: MedlinePlus