NF-κB对Cbl-b的调控及其在T细胞活化中作用的研究

The signaling pathway from TCR to T cell activation is a hot topic of transplant immunology. NF-κB and Cbl-b are both important signaling molecules in T cell activation. However, the regulation between them has not been clearly clarified. It has been demonstrated that Cbl-b can negatively regulate NF-κB translocation and T cell activation. Up to now, there is no report on the regulation of NF-κB on Cbl-b. Our preliminary study found that NF-κB can negatively regulate the transcription of Cbl-b. So, we speculate that this regulation may also be involved in T cell activation, which is an important signaling pathway in T cell activation. In order to evaluate the effects of regulation of NF-κB on Cbl-b in T cell activation, we will employ two special techniques to block the regulation of NF-κB on Cbl-b in this research. One is RNA interference to decoy corepressors of NF-κB. The other is TALENs (transcription activator-like effector nucleases) to induce mutation of the binding site of NF-κB in the promoter region of Cbl-b. This research will improve the signaling network of T cell activation from a new perspective and provide a new target for the treatment of transplant rejection.

T细胞活化信号通路的研究一直是移植免疫研究的热点。NF-κB和Cbl-b均是T细胞活化关键信号分子，二者间的调控关系尚未阐明，有研究发现Cbl-b可抑制NF-κB激活，阻断T细胞活化。但NF-κB对Cbl-b的表达有何影响，迄今国内外尚无报道。我们前期研究发现，在CHO细胞NF-κBp65可以负调控Cbl-b基因转录。据此我们推测这一调控机制也参与T细胞活化，这可能是T细胞活化过程中一个关键的信号通路。为证实这一假说，我们拟采用RNA干扰技术沉默相应NF-κBp65共抑制因子，TALENs的靶向基因修饰技术靶向诱导与NF-κBp65特异结合的Cbl-b启动子系列突变，阻断NF-κBp65 对Cbl-b的负调控作用，从细胞和整体水平评价NF-κBp65负调控Cbl-b表达在T细胞活化过程中的作用。本课题将从新的视角完善T细胞活化信号通路，并为移植免疫排斥的防治提供新的靶点。

NF-κB和Cbl-b均是T细胞活化关键信号分子，二者间的调控关系尚未阐明。我们前期体外研究发现NF-κB可结合于Cbl-b启动子序列并负调控其表达。本研究进一步利用双基因改造技术，在体内整体水平探索NF-κB与Cbl-b的调控关系。结果发现：（1）当将IκBαΔN-Tg小鼠的Cbl-b基因敲除后，能使耐受的移植心脏被排斥。（2）同IκBαΔN-Tg小鼠相比，双基因改造IκBαΔN-Tg/Cbl-b-/-小鼠在心脏移植后T细胞活化程度显著减低。（3）Cbl-b启动子突变影响NF-κB对Cbl-b启动子诱发的蛋白表达的调控。这些研究结果提示，Cbl-b是NF-κB的下游基因，而且控制着T细胞的活化和移植排斥反应的发生。结合前期体外实验结果，表明：NF-κB通过下调Cbl-b调控T细胞活化。