MOLECULAR MECHANISM OF T LYMPHOCYTE ACTIVATION

该实验室致力于确定细胞的作用， 癌基因Cbl在T淋巴细胞活化中的作用。 虽然Cbl的功能是 未知，它是酪氨酸磷酸化的主要目标后， 多种受体（包括T细胞受体（TCR））的结合， 这表明Cbl参与了信号转导途径， 由不同的受体类型触发。此外，Cbl还与 许多蛋白质被认为在调节 受体介导的激活事件。在T细胞中，我们发现， Cbl的过表达显著降低了TCR的活性 交联以激活AP 1（RAS）和NF-AT（PLC/Ca++） 激活途径。 在AP 1激活的情况下，Cbl似乎是 作用于Ras本身的上游，可能是通过调节 Grb 2与Sos（或其他胍交换因子）。尽管CBL在- 表达导致TCR后NF-AT活化显著降低， 交联时，它对NF-AT活性没有影响， 用PMA和离子霉素或活化形式的 Ca++敏感磷酸酶，钙调神经磷酸酶，这表明Cbl也可能 调节PLC g1的激活。最近的研究表明， 过表达的Cbl在产生 TCR交联后的磷酸肌醇磷酸（IP 3），表明PLCg 1 活动确实减少了。激活诱导的酪氨酸磷酸化 在Cbl过表达细胞中PLCg 1也减少。这进一步表明 Cbl参与调控PLCg 1的激活， 受体介导的T细胞活化。 因为Cbl可以调节 RAS和PLCg 1/ Ca++激活途径，我们认为这些数据表明， Cbl作为一个“复杂的”衔接分子。在这个角色中，Cbl可以通过以下方式行事： 预先组装效应分子和衔接分子的有序复合物， TCR活化，并促进有效的信号转导后， 受体结合在未激活的细胞中组装这种复合物时， Cbl也可能调节特定效应物与此的相互作用。 复杂. Cbl过表达通过破坏 活化复合物的生理化学计量。我们 目前正在产生Cbl截短和缺失突变体， 用于解决Cbl的哪些区域需要用于调节 每一个途径，并确定Cbl相关的分子， 对TCR介导的RAS和PLCg 1激活都至关重要。

The laboratory is concened with identifying the role of the cellular oncogene, Cbl, in T lymphocyte activation. While the function of Cbl is unknown, it is a major target of tyrosine phosphorylation after the engagement of multiple receptors (including the T cell receptor (TCR)), suggesting that Cbl is involved in the signal transduction pathways triggered by distinct receptor types. In addition, Cbl associates with numerous proteins which are thought to be active in the regulation of receptor mediated activation events. In T cells we have found that over-expression of Cbl significantly reduced the abiltiy of TCR cross-linking to activate both the AP1 (RAS) and the NF-AT (PLC/Ca++) activation pathways. In the case of AP1 activation, Cbl appears to be acting upstream of Ras itself, possibly by regulating the association of Grb2 with Sos (or other guanidine exchange factors). Although Cbl over- expression led to a significant decrease in NF-AT activation after TCR cross-linking, it had no affect on NF-AT activity when cells were stimulated with PMA and ionomycin or an activated form of the Ca++-sensitive phosphatase, calcineurin, suggesting that Cbl may also regulate PLCg1 activation. Recent studies indicate that cells that over-express Cbl have a significant decrease in the generation of phosphoinositol phosphates (IP3) after TCR-cross-linking suggesting PLCg1 activity is indeed reduced. Activation induced tyrosine phosphorylation of PLCg1 in also reduced in Cbl over-expressing cells. This further suggests that Cbl participates in the regulation of PLCg1 activation during receptor-mediated T cell activation. Because Cbl can regulate both the RAS and PLCg1/ Ca++ activation pathways we feel these data suggest that Cbl acts as a "complex" adapter molecule. In this role, Cbl may act by pre-assembling an ordered complex of effector and adapter molecules prior to TCR activation, and facilitating efficient signal transduction after receptor engagement. In assembling such a complex in unactivated cells, Cbl may also egulate the interaction of specific effectors with this complex. Cbl over-expression negatively affects activation by disrupting the pyhsiologic stoichiometry of the activation complexes. We are currently generating Cbl truncation and deletion mutants which will be used to address which regions of Cbl are required for the regulation of each pathway and to identify the Cbl-associated molecules which are critical for both TCR-mediated RAS and PLCg1 activation.