MOLECULAR MECHANISM OF T LYMPHOCYTE ACTIVATION

实验室致力于确定细胞的作用。 癌基因Cb1在T淋巴细胞活化中的作用而Cbl的功能是 未知，它是酪氨酸磷酸化的主要靶点 多种受体(包括T细胞受体(TCR))的参与， 提示Cb1参与了信号转导途径。 由不同的受体类型触发。此外，Cbl还与 许多蛋白质被认为是活跃的调节 受体介导的激活事件。在T细胞中，我们发现 Cb1的过表达显著降低了TCR的表达能力 激活AP1(RAS)和NF-AT(PLC/Ca)的交联性 激活途径。在AP1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 Grb2与SOS(或其他胍交换因子)。尽管Cbl结束了- TCR后核因子-AT的表达显著降低 交联剂对细胞内的核因子-AT活性无影响 用PMA和离子霉素或其激活形式刺激 CA敏感的磷酸酶，钙调神经磷酸酶，提示Cb1也可能 调节PLCG1的激活。最近的研究表明，细胞 过度表达的Cbl显著减少了 TCR交联后的磷酸肌醇磷酸盐(IP3)提示PLCG1 活动确实减少了。激活诱导的酪氨酸磷酸化 PLCG1在Cb1过表达细胞中也减少。这进一步表明 Cb1参与PLCG1激活的调节 受体介导的T细胞活化。因为Cbl既可以监管 RAS和PLCG1/Ca激活途径我们认为这些数据表明 CBL是一种“复杂的”适配分子。在这个角色中，Cbl可以通过 预先组装效应器和接头分子的有序复合体 TCR激活，并促进有效的信号转导 感受器接触。在未激活的细胞中组装这样的复合体时， CBL还可以调节特定效应器与此的相互作用 很复杂。CBL的过度表达通过干扰对激活产生负面影响 活化络合物的生理化学计量学。我们是 目前正在生成Cbl截断和删除突变体，这将是 用于说明Cbl的哪些区域是调节所需的 每个途径，并鉴定Cbl相关分子，这些分子是 对TCR介导的RAS和PLCG1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.