【作者】 陈伟；

【导师】 肖能明；

【作者基本信息】 厦门大学 ， 生物化学与分子生物学， 2021， 博士

【摘要】 转录抑制因子Zeb1广泛表达于各种免疫细胞中,通过调节转录网络在免疫细胞的发育、分化和功能方面发挥着关键的作用。TCR信号的起始、T细胞活化和扩增是效应T细胞和记忆T细胞的分化和功能发挥的关键前提,更重要的是T细胞扩增阶段是T细胞死亡拮抗的过程。本论文研究发现Zeb1并不为T细胞活化和增殖所必需,但是Zeb1通过阻止T细胞凋亡以维持T细胞的扩增,从而为抗病毒和抗肿瘤T细胞免疫所必需。机制方面,首先Zeb1经典靶基因Cdh1的遗传删除并不能改善缺失Zeb1造成的T细胞扩增的缺陷。其次,芳香族化合物受体AhR的转录也可以直接被Zeb1抑制;过表达AhR能够充分诱导T细胞死亡,同时内源性配体FICZ和ITE或者外源性配体Bap和βNF激活AhR也可以加速EL4-AhR细胞的死亡。相对应地,Ido抑制剂D-1MT、L-1MT和D/L-1MT或者AhR抑制剂PDM2在体外也可以完全阻止T细胞的死亡。但是,遗传水平删除AhR不能阻止因缺失Zeb1导致T细胞的死亡。最后,我们也发现敲除Zeb1的T细胞存在DNA损伤诱导的Chk1/Caspase-2凋亡通路和Caspase-8介导的外源凋亡通路。其中,Chk1的活性被上游多个分子所调节,其中包含Kdm1a、Hus1和Claspin;Zeb1缺失的T细胞中Kdm1a蛋白水平异常升高和Hus1以及Claspin蛋白水平异常下调均能促使Chk1的S345位点磷酸化减少,进而削减Chk1的活性,导致Caspase-2介导的凋亡通路被激活。此外AhR抑制剂PDM2可以完全抑制T细胞凋亡信号的发生,抑制剂PDM2或许可以成为肿瘤免疫治疗的新的小分子化合物候选。这些结果表明Zeb1可以通过协同转录网络抑制多个凋亡通路以维持T细胞存活,继而促进T细胞的扩增,这为抗病毒和抗肿瘤T细胞免疫是至关重要的。更多还原

【Abstract】 The transcription repressor Zebl is ubiquitously expressed in various immune cells and plays a critical role in development,differentiation and function of immune cells.Initiation of TCR signaling,T cell activation and expansion are nessesary for the differentiation and function of effector and memory T cells.More importamtly,T cell is resistant to apoptosis during T cells expansion.In this study,we found Zebl was dispensible for activation and proliferation of T cells,but Zeb1 is intrinsically required for effective antiviral and antitumor T cell immunity through preventing T cells from apoptosis to maintain T cell expansion.Firstly,genetic ablation of Cdhl,a well-known target gene of Zebl,did not rectify the defective expansion of Zeb1-deficient T cells.Secondly,AhR was directly repressed by Zebl in T cells,and its ectopic expression accelelated antigen-specific T cell death in vivo.Moreover,AhR activation by endogenous ligands(FICZ or ITE)or exogenous ligands(Bap or βNF)significantly impeded expansion of AhR-overexpressing EL4 cells in vitro.In contrast,cell death of Zeb1-deficient T cells was ameliorated by Ido inhibitors(D-1MT,L-1MT or D/L-1MT)or AhR antagonist PDM2 in vitro.However,neither AhR deficiency nor Casp8/Ripk3 double deficiency restored cell death in Zeb1-deficient T cells.Furthermore,in addition to Caspase-8-mediated cell apoptosis,Zeb1-deficient T cells also underwent DNA damage-induced Caspase-2-mediated cell apoptosis.Zeb1 deficiency in T cells resulted in accumulation of DNA damage,up-regulation of Kdm1a expression and downregualtion of Husl and Claspin expression,which cooparatively attenuated phosphorylation and activation of Chkl and subsequently led to Caspase-2-mediated cell apoptosis.Last,apoptosis pathways in T cells lacking Zebl were completely blocked by PDM2 in vitro,PDM2 might be a novel candidate of small compound in tumor immunotherapy.Thus,our results indicate that Zeb1 sustains T cell survival and expansion by restricting multiple apoptosis pathways,including both Caspase-2-and Caspase-8-mediated cell apoptosis,which is crucial to protect T cells against virus infection and tumorigenesis.更多还原