The human papillomavirus 16 E6 protein binds to Fas-associated death domain and protects cells from Fas-triggered apoptosis
The human papillomavirus 16 E6 protein binds to Fas-associated death domain and protects cells from Fas-triggered apoptosis
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DOI：
10.1074/jbc.m401172200
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发表时间：
2004-06-11
影响因子：
4.8
通讯作者：
Duerkesen-Hughes, PJ
中科院分区：
文献类型：
--
作者：
Filippova, M;Parkhurst, L;Duerkesen-Hughes, PJ
人类乳头瘤病毒(HPV)的高危株，如HPV16，会导致人类宫颈癌。HPV16的E6蛋白介导了抑癌基因P53的快速降解，尽管这不是E6的唯一功能，也不能完全解释其转化潜力。我们实验室以前的工作已经证明，E6可以通过与肿瘤坏死因子R1的C末端结合，从而阻断细胞凋亡信号转导，从而保护细胞免受肿瘤坏死因子诱导的细胞凋亡。在这项研究中，E6也被证明可以保护细胞免受通过Fas途径诱导的细胞凋亡。此外，使用可诱导的E6表达系统证明了这种保护是剂量依赖的，更高水平的E6导致更好的保护。虽然E6抑制caspase3和caspase8的激活，但它不影响通过线粒体途径的凋亡信号。然后用哺乳动物双杂交和体外下拉实验证明E6直接与Fas相关死亡结构域(FADD)的死亡效应域结合，缺失和定点突变使E6结合位点定位到FADD死亡效应域的N端。E6以两种形式产生：类似于16 kDa的全长版本和大约一半大小的较小版本，对应于全长蛋白质的N-末端一半。下拉和功能分析表明，E6的全长版本，而不是小版本，能够与FADD结合，并保护细胞免受Fas诱导的细胞凋亡。此外，与E6结合会导致FADD的降解，细胞内FADD的损失与E6的表达量成正比。这些结果支持一个模型，在该模型中，E6介导的FADD降解阻止了凋亡信号通过Fas途径的传递。
High risk strains of human papillomavirus (HPV), such as HPV 16, cause human cervical carcinoma. The E6 protein of HPV 16 mediates the rapid degradation of the tumor suppressor p53, although this is not the only function of E6 and cannot completely explain its transforming potential. Previous work in our laboratory has demonstrated that E6 can protect cells from tumor necrosis factor-induced apoptosis by binding to the C-terminal end of tumor necrosis factor R1, thus blocking apoptotic signal transduction. In this study, E6 was shown to also protect cells from apoptosis induced via the Fas pathway. Furthermore, use of an inducible E6 expression system demonstrated that this protection is dose-dependent, with higher levels of E6 leading to greater protection. Although E6 suppresses activation of both caspase 3 and caspase 8, it does not affect apoptotic signaling through the mitochondrial pathway. Mammalian two-hybrid and in vitro pull-down assays were then used to demonstrate that E6 binds directly to the death effector domain of Fas-associated death domain (FADD), with deletion and site-directed mutants enabling the localization of the E6-binding site to the N-terminal end of the FADD death effector domain. E6 is produced in two forms as follows: a full-length version of similar to16 kDa and a smaller version of about half that size corresponding to the N-terminal half of the full-length protein. Pull-down and functional assays demonstrated that the full-length version, but not the small version of E6, was able to bind to FADD and to protect cells from Fas-induced apoptosis. In addition, binding to E6 leads to degradation of FADD, with the loss of cellular FADD proportional to the amount of E6 expressed. These results support a model in which E6-mediated degradation of FADD prevents transmission of apoptotic signals via the Fas pathway.