Mechanism of Fas Signaling Regulation by Human Herpesvirus 8 K1 Oncoprotein
Mechanism of Fas Signaling Regulation by Human Herpesvirus 8 K1 Oncoprotein
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DOI：
10.1093/jnci/djn516
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发表时间：
2009-03-18
影响因子：
10.3
通讯作者：
Samaniego, Felipe
中科院分区：
文献类型：
--
作者：
Berkova, Zuzana;Wang, Shu;Samaniego, Felipe
疱疹病毒8型（HHV-8）癌蛋白K1与Fas死亡受体介导的淋巴细胞增殖和细胞凋亡抑制有关。K1在转基因小鼠中的表达诱导淋巴组织细胞和淋巴瘤的积累。方法为了研究K1和Fas如何相互作用以抑制凋亡，在表达野生型K1或K1 IG结构域缺失突变体的人胚肾（HEK）和淋巴瘤（BJAB）细胞中研究K1-Fas结合，并用Fas配体（FasL）或激动性Fas抗体处理，免疫沉淀和蛋白质印迹分析。在来自用空载体与用表达野生型K1或K1 IG缺失突变体的质粒转染并用激动性Fas抗体处理7小时的小鼠的肝样品中比较胱天蛋白酶-3的切割和细胞凋亡。这些研究采用免疫组织化学染色和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法。结果转染HEK和BJAB细胞的免疫沉淀和Western印迹分析显示，野生型K1，但不是Ig缺失的K1结合Fas和阻止Fas激活的FasL或由激动性Fas抗体。用野生型K1转染的小鼠（7/10）比用空载体转染的小鼠（3/13）或K1 IG缺失突变体转染的小鼠（0/6）在用激动性Fas抗体处理后存活更多。与载体转染小鼠相比，野生型K1转染小鼠的肝脏含有较少的caspase-3裂解细胞（87.6% vs 58.0%，差异= 29.6%，95%置信区间[CI] = 19.2%至40.0%; P=.003）和更少的凋亡细胞（83.7%vs 34.2%，difference = 49.5%，95%CI =39.8%至59.2%; P=.003）。结论K1通过其Ig样结构域直接与Fas结合并阻止FasL的结合而阻断Fas信号传导。癌细胞对Fas介导的凋亡的相对抗性可能是由于含有IG结构域的蛋白抑制Fas。
Background Herpesvirus 8 (HHV-8) oncoprotein K1 is linked to lymphoproliferation and suppression of apoptosis mediated by the Fas death receptor. Expression of K1 in transgenic mice induces accumulation of lymphoid tissue cells and lymphoma.Methods To examine how K1 and Fas interact to suppress apoptosis, K1-Fas binding was studied in human embryonic kidney (HEK) and lymphoma (BJAB) cells that expressed wild-type K1 or a K1 Ig domain deletion mutant and were treated with Fas ligand (FasL) or an agonistic Fas antibody, using immunoprecipitation and Western blot analysis. Cleavage of caspase-3 and apoptosis was compared in liver samples from mice that were transfected with empty vector vs with plasmids expressing wild-type K1 or a K1 Ig deletion mutant and treated with agonistic Fas antibody for 7 hours. These studies used immunohistochemical staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. All statistical tests were two-sided.Results Immunoprecipitation and Western blot analysis of transfected HEK and BJAB cells revealed that wild-type K1 but not Ig-deleted K1 binds to Fas and prevents Fas activation by FasL or by an agonistic Fas antibody. More mice that were transfected with wild-type K1 (7 of 10) than mice transfected with empty vector (3 of 13) or the K1 Ig deletion mutant (0 of 6) survived treatment with the agonistic Fas antibody. Compared with vector-transfected mice, livers of wild-type K1-transfected mice contained fewer cells in which caspase-3 was cleaved (87.6% vs 58.0%, difference = 29.6%, 95% confidence interval [CI] = 19.2% to 40.0%; P=.003) and fewer apoptotic cells (83.7% vs 34.2%, difference = 49.5%, 95% CI =39.8% to 59.2%; P=.003).Conclusions K1 blocks Fas signaling by directly binding to Fas through the Ig-like domain of K1 and preventing binding of FasL. The relative resistance of cancer cells to Fas-mediated apoptosis may be due to the inhibition of Fas by Ig domain-containing proteins.