Accumulation of Heterochromatin Components on the Terminal Repeat Sequence of Kaposi's Sarcoma-Associated Herpesvirus Mediated by the Latency-Associated Nuclear Antigen
Accumulation of Heterochromatin Components on the Terminal Repeat Sequence of Kaposi's Sarcoma-Associated Herpesvirus Mediated by the Latency-Associated Nuclear Antigen
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DOI：
10.1128/jvi.78.14.7299-7310.2004
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发表时间：
2004-07
影响因子：
5.4
通讯作者：
S. Sakakibara;K. Ueda;K. Nishimura;E. Do;Eriko Ohsaki;T. Okuno;K. Yamanishi
中科院分区：
文献类型：
--
作者：
S. Sakakibara;K. Ueda;K. Nishimura;E. Do;Eriko Ohsaki;T. Okuno;K. Yamanishi
卡波济肉瘤相关疱疹病毒（KSHV）潜伏感染时，其160 kb环状游离DNA在宿主细胞核内复制并维持。KSHV潜伏相关核抗原（拉娜）是维持病毒潜伏期的关键因子。拉娜与病毒基因组的末端重复（TR）DNA结合，导致其定位于细胞核中的特定点结构。在这样的感染细胞中，病毒基因的表达受到一种机制的限制，这种机制目前还不清楚。在这里，我们发现，拉娜相互作用与SUV 39 H1组蛋白甲基转移酶，异染色质形成的关键组成部分，通过使用DNA下拉测定与生物素化的DNA片段，含有LANA特异性结合序列和麦芽糖结合蛋白下拉测定。拉娜在未感染细胞染色体上的扩散定位随着含有大部分TR区域的细菌人工染色体的引入而变为点状的，并且SUV 39 H1清楚地与LANA相关的点共定位。因此，KSHV感染细胞中的拉娜病灶似乎包括SUV 39 H1以及异染色质蛋白1。此外，染色质免疫沉淀试验显示，TR和开放阅读框（ORF）K1和ORF 50/RTA基因，但不是ORF 73/拉娜基因，在KSHV潜伏期异染色质内。综上所述，这些观察结果表明，拉娜招募异染色质组分到病毒基因组中，这可能导致病毒潜伏期的建立并控制转录程序。
ABSTRACT In the latent infection of Kaposi's sarcoma-associated herpesvirus (KSHV), its 160-kb circularized episomal DNA is replicated and maintained in the host nucleus. KSHV latency-associated nuclear antigen (LANA) is a key factor for maintaining viral latency. LANA binds to the terminal repeat (TR) DNA of the viral genome, leading to its localization to specific dot structures in the nucleus. In such an infected cell, the expression of the viral genes is restricted by a mechanism that is still unclear. Here, we found that LANA interacts with SUV39H1 histone methyltransferase, a key component of heterochromatin formation, as determined by use of a DNA pull-down assay with a biotinylated DNA fragment that contained a LANA-specific binding sequence and a maltose-binding protein pull-down assay. The diffuse localization of LANA on the chromosomes of uninfected cells changed to a punctate one with the introduction of a bacterial artificial chromosome containing most of the TR region, and SUV39H1 clearly colocalized with the LANA-associated dots. Thus, the LANA foci in KSHV-infected cells seemed to include SUV39H1 as well as heterochromatin protein 1. Furthermore, a chromatin immunoprecipitation assay revealed that the TR and the open reading frame (ORF) K1 and ORF50/RTA genes, but not the ORF73/LANA gene, lay within the heterochromatin during KSHV latency. Taken together, these observations indicate that LANA recruits heterochromatin components to the viral genome, which may lead to the establishment of viral latency and govern the transcription program.