The molecular mechanism of TCR repertoire generation and coreceptor expression

为了研究TCR库生成的分子机制，我们分析了调节TCR、CD4和_cd8共受体表达的相关分子。在基因操纵小鼠的使用中，我们证明了以下几点。(1)在DP期随机升高的TCR表达在没有PS的情况下被TCR- mhc相互作用下调，但在没有适当的TCR介导信号的情况下保持其密度。这一结论与之前的假设不一致，即DP细胞上TCR的表达本来是低的，但当DP细胞接受TCR介导的PS信号时，TCR的表达被诱导升高，而没有TCR信号时，TCR的表达不增加。然而，我们的研究结果有助于解决先前假设中提出的问题，即低TCR如何以低亲和度识别MHC分子，从而可能正向选择DP细胞，导致DP细胞激活以高TCR表达。(2) DP细胞上的CD45与TCR表达的下调密切相关，而TCR介导的信号能够克服这种下调。(3)我们探索了体内胸腺细胞胞内底物酪氨酸磷酸化的检测，发现ZAP-70在CD8SP细胞系的PS过程中高度磷酸化，而在CD4SP细胞系中则没有磷酸化。这一结果与最近报道的一例人类ZAP-70异常患者的观察结果一致，该患者未出现CD8SP细胞，但发现了CD4SP细胞，但数量较少。这些发现提示了DP胸腺细胞CD4和CD8下调的不对称机制。(4) CD4成熟T细胞通过非mhc相关调控进一步功能发育为Th1和Th2细胞。

To study molecular mechanism of positive selection (PS) process for generating TCR repertoire, we analyzed related molecules for regulating expression of TCR and CD4 and_CD8 co-receptor. In use of gene manipulating mice, we proved followings.(1) TCR expression which increased randomly at DP stage was down-regulated without PS by TCR-MHC interaction but maintained its density without an appropriate TCR mediated signal. This conclusion is not consistent with the previous hypothesis that TCR expression on DP cells is originally low but it is induced to raise up when DP cells receive TCR mediated PS signal while TCR expression is not increased without TCR signal. However, our findings is helpful to resolve the question raised in the previous hypothesis how low TCR can recognize MHC molecules with low avidity which may positively select DP cells, resulting in DP cells activation for high TCR expression. (2) CD45 on DP cells was closely involved in down-regulation of TCR expression, which was overcome by TCR mediated signal. (3) We explored to detect tyrosine phosphorylation of intracellular substrates of thymocytes in the in vivo condition, and demonstrated that ZAP-70 is highly phosphorylated in PS process for CD8SP generation but not for CD4SP cell lineage. This result is consistent with the recent observation reported in a human patient possessing with abnormal ZAP-70, in which CD8SP cells did not develop but CD4SP cells were found though their number was small. These findings suggest an asymmetric mechanism for down regulation of CD4 and CD8 in DP thymocytes. (4) CD4 mature T cells are induced further functional development into Th1 and Th2 cells by non-MHC related regulation.