Involvement of p21ras distinguishes positive and negative selection in thymocytes.

Abstract

Small molecular weight GTP binding proteins of the ras family have been implicated in signal transduction from the T cell antigen receptor (TCR). To test the importance of p21ras in the control of thymocyte development, we generated mice expressing a dominant‐negative p21ras protein (H‐rasN17) in T lineage cells under the control of the lck proximal promoter. Proliferation of thymocytes from lck‐H‐rasN17 mice in response to TCR stimulation was nearly completely blocked, confirming the importance of p21ras in mediating TCR‐derived signals in mature CD4+8‐ or CD8+4‐ thymocytes. In contrast, some TCR‐derived signals proceeded unimpaired in the CD4+8+ thymocytes of mice expressing dominant‐negative p21ras. Analysis of thymocyte development in mice made doubly transgenic for the H‐Y‐specific TCR and lck‐H‐rasN17 demonstrated that antigen‐specific negative selection occurs normally in the presence of p21H‐rasN17. Superantigen‐induced negative selection in vivo also proceeded unhindered in H‐rasN17 thymocytes. In contrast, positive selection of thymocytes in the H‐Y mice was severely compromised by the presence of p21H‐rasN17. These observations demonstrate that positive and negative selection, two conceptually antithetical consequences of TCR stimulation, are biochemically distinguishable.

B‐Raf‐mediated signaling pathway regulates T cell development

“…Transgenic expression of dominantnegative or constitutively active forms of Ras, Raf or MEK1 all revealed that the MAPK pathway is implicated in positive selection at the CD4 + CD8 + DP stage [6][7][8][9]. Consistent with these reports, ERK activation is involved in the most critical events during TCR signaling and plays a crucial role in positive selection [4,5].…”

“…The defect of ERK activation was not observed in B-Raf -/-thymocytes treated with PMA, which provoked substantial activation of the Raf-1-ERK pathway. It is interesting to note that thymocytes expressing dominant-interfering mutants of Raf and Ras also exhibited DP arrest [6][7][8] a) Thymi from control B-Raf +/+ chimeras (n=4) or B-Raf -/-chimeras (n=4) were harvested 8-12 wk after transfer, and cells were stained with the indicated subset marker. b) p values were calculated for frequencies using a two-tailed Student's t-test, which was considered significant at p<0.05. c) Each value represents the frequency of the indicated population within total thymocytes or the CD4 + CD8 + DP population.…”

“…After DP cells undergo positive selection, the expression of TCR increases, and thus TCR + DP and CD4 + or CD8 + SP cells are expected to be positively selected [3,7,8]. We therefore examined the TCR expression on B-Raf +/+ and B-Raf -/-thymocytes.…”

Transgenic overexpression of the Caspase‐8 inhibitor FLIPshort leads to impaired T cell proliferation and an increased memory T cell pool after staphylococcal enterotoxin B injection

“…Human FLAG-tagged c-FLIP S cDNA was cloned blunt-ended into the Bam HI site of the p1017 transgenic vector [46]. This vector contains the proximal (3.2 kb) lck promoter and, downstream of the cDNA insertion site, 2.1 kb of genomic sequence of the human growth hormone gene (hGH).…”

Duration and Strength of Extracellular Signal-Regulated Kinase Signals Are Altered During Positive Versus Negative Thymocyte Selection

“…Studies have shown that mice overexpressing dominant negative components of the Ras (MAP) kinase signaling pathway (Ras/raf-1/MAP kinase kinase (MEK)) exhibit partially blocked positive selection (5)(6)(7)(8) and extracellular signal-regulated kinase (ERK) 1-deficient thymocytes show impaired positive selection (9). A selective role for ERK in positive selection was also shown in mice expressing a mutation in the TCR ␣-chain-connecting peptide (␣CPM).…”