Lyu Yuanzhi, Nakano Kazushi, Davis Ryan R, Tepper Clifford G, Campbell Mel, Izumiya Yoshihiro
Department of Dermatology, School of Medicine, University of California, Davis, Sacramento, California, USA.
Department of Biochemistry and Molecular Medicine, School of Medicine, University of California, Davis, Sacramento, California, USA.
J Virol. 2017 Oct 13;91(21). doi: 10.1128/JVI.00980-17. Print 2017 Nov 1.
Bivalent histone modifications are defined as repressive and activating epigenetic marks that simultaneously decorate the same genomic region. The H3K27me3 mark silences gene expression, while the H3K4me3 mark prevents the region from becoming permanently silenced and prepares the domain for activation when needed. Specific regions of Kaposi's sarcoma-associated herpesvirus (KSHV) latent episomes are poised to be activated by the KSHV replication and transcription activator (K-Rta). How KSHV episomes are prepared such that they maintain latent infection and switch to lytic replication by K-Rta remains unclear. K-Rta transactivation activity requires a protein degradation function; thus, we hypothesized that identification of cellular substrates of K-Rta may provide insight into the maintenance of KSHV latent infection and the switch to lytic replication. Here we show that a zinc finger protein, ZIC2, a key regulator for central nervous system development, is a substrate of K-Rta and is responsible for maintaining latency. K-Rta directly interacted with ZIC2 and functioned as an E3 ligase to ubiquitinate ZIC2. ZIC2 localized at immediate early and early gene cluster regions of the KSHV genome and contributed to tethering of polycomb repressive complex 2 through physical interaction, thus maintaining H3K27me3 marks at the K-Rta promoter. Accordingly, depletion of ZIC2 shifted the balance of bivalent histone modifications toward more active forms and induced KSHV reactivation in naturally infected cells. We suggest that ZIC2 turnover by K-Rta is a strategy employed by KSHV to favor the transition from latency to lytic replication. Posttranslational histone modifications regulate the accessibility of transcriptional factors to DNA; thus, they have profound effects on gene expression (e.g., viral reactivation). KSHV episomes are known to possess bivalent chromatin domains. How such KSHV chromatin domains are maintained to be reactivatable by K-Rta remains unclear. We found that ZIC2, a transcriptional factor essential for stem cell pluripotency, plays a role in maintaining KSHV latent infection in naturally infected cells. We found that ZIC2 degradation by K-Rta shifts bivalent histone marks to a more active configuration, leading to KSHV reactivation. ZIC2 interacts with and maintains polycomb repressor complex 2 at the K-Rta promoter. Our findings uncover (i) a mechanism utilized by KSHV to maintain latent infection, (ii) a latency-lytic cycle switch operated by K-Rta, and (iii) a molecular mechanism of ZIC2-mediated local histone modification.
二价组蛋白修饰被定义为同时修饰同一基因组区域的抑制性和激活性表观遗传标记。H3K27me3标记使基因表达沉默，而H3K4me3标记可防止该区域永久沉默，并在需要时为激活该结构域做好准备。卡波西肉瘤相关疱疹病毒（KSHV）潜伏附加体的特定区域准备好被KSHV复制和转录激活因子（K-Rta）激活。KSHV附加体如何做好准备以维持潜伏感染并通过K-Rta切换到裂解复制仍不清楚。K-Rta的反式激活活性需要蛋白质降解功能；因此，我们推测鉴定K-Rta的细胞底物可能有助于深入了解KSHV潜伏感染的维持以及向裂解复制的转变。在这里，我们表明锌指蛋白ZIC2是中枢神经系统发育的关键调节因子，是K-Rta的底物，并负责维持潜伏状态。K-Rta直接与ZIC2相互作用，并作为E3连接酶使ZIC2泛素化。ZIC2定位于KSHV基因组的立即早期和早期基因簇区域，并通过物理相互作用有助于多梳抑制复合物2的拴系，从而在K-Rta启动子处维持H3K27me3标记。因此，ZIC2的缺失使二价组蛋白修饰的平衡向更活跃的形式转变，并在自然感染的细胞中诱导KSHV重新激活。我们认为，K-Rta介导的ZIC2周转是KSHV采用的一种策略，有利于从潜伏状态向裂解复制的转变。翻译后组蛋白修饰调节转录因子对DNA的可及性；因此，它们对基因表达（如病毒重新激活）有深远影响。已知KSHV附加体具有二价染色质结构域。这种KSHV染色质结构域如何被维持以便被K-Rta重新激活仍不清楚。我们发现ZIC2是干细胞多能性所必需的转录因子，在自然感染的细胞中对维持KSHV潜伏感染起作用。我们发现K-Rta介导ZIC2的降解会使二价组蛋白标记转变为更活跃的构型，导致KSHV重新激活。ZIC2在K-Rta启动子处与多梳抑制复合物2相互作用并维持其存在。我们的研究结果揭示了（i）KSHV用于维持潜伏感染的机制，（ii）由K-Rta操作的潜伏-裂解周期转换，以及（iii）ZIC2介导的局部组蛋白修饰的分子机制。
