Division of Molecular and Experimental Surgery, Department of Surgery, University Medical Center Erlangen, Friedrich-Alexander-University Erlangen-Nürnberg, Erlangen, Germany.
Department of Mathematical Sciences, University of Essex, Colchester, UK.
Oncogene. 2015 Jan 29;34(5):639-49. doi: 10.1038/onc.2013.595. Epub 2014 Jan 27.
The tumor suppressor p53 is a central regulatory molecule of apoptosis and is commonly mutated in tumors. Kaposi's sarcoma-associated herpesvirus (KSHV)-related malignancies express wild-type p53. Accordingly, KSHV encodes proteins that counteract the cell death-inducing effects of p53. Here, the effects of all KSHV genes on the p53 signaling pathway were systematically analyzed using the reversely transfected cell microarray technology. With this approach we detected eight KSHV-encoded genes with potent p53 inhibiting activity in addition to the previously described inhibitory effects of KSHV genes ORF50, K10 and K10.5. Interestingly, the three most potent newly identified inhibitors were KSHV structural proteins, namely ORF22 (glycoprotein H), ORF25 (major capsid protein) and ORF64 (tegument protein). Validation of these results with a classical transfection approach showed that these proteins inhibited p53 signaling in a dose-dependent manner and that this effect could be reversed by small interfering RNA-mediated knockdown of the respective viral gene. All three genes inhibited p53-mediated apoptosis in response to Nutlin-3 treatment in non-infected and KSHV-infected cells. Addressing putative mechanisms, we could show that these proteins could also inhibit the transactivation of the promoters of apoptotic mediators of p53 such as BAX and PIG3. Altogether, we demonstrate for the first time that structural proteins of KSHV can counteract p53-induced apoptosis. These proteins are expressed in the late lytic phase of the viral life cycle and are incorporated into the KSHV virion. Accordingly, these genes may inhibit cell death in the productive and in the early entrance phase of KSHV infection.
肿瘤抑制因子 p53 是细胞凋亡的核心调节分子，通常在肿瘤中发生突变。卡波西肉瘤相关疱疹病毒（KSHV）相关恶性肿瘤表达野生型 p53。相应地，KSHV 编码的蛋白可以抵消 p53 诱导细胞死亡的作用。在这里，我们使用反向转染细胞微阵列技术系统地分析了所有 KSHV 基因对 p53 信号通路的影响。通过这种方法，除了先前描述的 KSHV 基因 ORF50、K10 和 K10.5 的抑制作用外，我们还检测到了 8 种具有强烈 p53 抑制活性的 KSHV 编码基因。有趣的是，新鉴定的三种最有效的抑制剂是 KSHV 结构蛋白，即 ORF22（糖蛋白 H）、ORF25（主要衣壳蛋白）和 ORF64（被膜蛋白）。用经典转染方法验证这些结果表明，这些蛋白以剂量依赖的方式抑制 p53 信号，并且可以通过各自病毒基因的小干扰 RNA 介导的敲低来逆转这种作用。这三种基因都可以抑制非感染和 KSHV 感染细胞中 Nutlin-3 处理后 p53 介导的细胞凋亡。在探讨潜在机制时，我们可以证明这些蛋白也可以抑制 p53 诱导的凋亡介质如 BAX 和 PIG3 的启动子的转录激活。总的来说，我们首次证明了 KSHV 的结构蛋白可以拮抗 p53 诱导的细胞凋亡。这些蛋白在病毒生命周期的晚期裂解期表达，并被整合到 KSHV 病毒粒子中。因此，这些基因可能在 KSHV 感染的有性和早期进入阶段抑制细胞死亡。
