MOE& MOH Key Laboratory of Medical Molecular Virology, School of Basic Medicine, Shanghai Medical College, Fudan University, Shanghai, People's Republic of China Department of Clinical Laboratory, Central Hospital of Taizhou City, Taizhou, People's Republic of China.
MOE& MOH Key Laboratory of Medical Molecular Virology, School of Basic Medicine, Shanghai Medical College, Fudan University, Shanghai, People's Republic of China.
J Virol. 2014 Jun;88(12):6873-84. doi: 10.1128/JVI.00283-14. Epub 2014 Apr 2.
Hypoxia-inducible factor 1α (HIF-1α) has been frequently implicated in many cancers as well as viral pathogenesis. Kaposi's sarcoma-associated herpesvirus (KSHV) is linked to several human malignancies. It can stabilize HIF-1α during latent infection and undergoes lytic replication in response to hypoxic stress. However, the mechanism by which KSHV controls its latent and lytic life cycle through the deregulation of HIF-1α is not fully understood. Our previous studies showed that the hypoxia-sensitive chromatin remodeler KAP1 was targeted by the KSHV-encoded latency-associated nuclear antigen (LANA) to repress expression of the major lytic replication and transcriptional activator (RTA). Here we further report that an RNA interference-based knockdown of KAP1 in KSHV-infected primary effusion lymphoma (PEL) cells disrupted viral episome stability and abrogated sub-G1/G1 arrest of the cell cycle while increasing the efficiency of KSHV lytic reactivation by hypoxia or using the chemical 12-O-tetradecanoylphorbol-13-acetate (TPA) or sodium butyrate (NaB). Moreover, KSHV genome-wide screening revealed that four hypoxia-responsive clusters have a high concurrence of both RBP-Jκ and HIF-1α binding sites (RBS+HRE) within the same gene promoter and are tightly associated with KAP1. Inhibition of KAP1 greatly enhanced the association of RBP-Jκ with the HIF-1α complex for driving RTA expression not only in normoxia but also in hypoxia. These results suggest that both KAP1 and the concurrence of RBS+HRE within the RTA promoter are essential for KSHV latency and hypoxia-induced lytic reactivation.
Kaposi's sarcoma-associated herpesvirus (KSHV), a DNA tumor virus, is an etiological agent linked to several human malignancies, including Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL). HIF-1α, a key hypoxia-inducible factor, is frequently elevated in KSHV latently infected tumor cells and contributes to KSHV lytic replication in hypoxia. The molecular mechanisms of how KSHV controls the latent and lytic life cycle through deregulating HIF-1α remain unclear. In this study, we found that inhibition of hypoxia-sensitive chromatin remodeler KAP1 in KSHV-infected PEL cells leads to a loss of viral genome and increases its sensitivity to hypoxic stress, leading to KSHV lytic reactivation. Importantly, we also found that four hypoxia-responsive clusters within the KSHV genome contain a high concurrence of RBP-Jκ (a key cellular regulator involved in Notch signaling) and HIF-1α binding sites. These sites are also tightly associated with KAP1. This discovery implies that KAP1, RBP-Jκ, and HIF-1α play an essential role in KSHV pathogenesis through subtle cross talk which is dependent on the oxygen levels in the infected cells.
缺氧诱导因子 1α（HIF-1α）在许多癌症以及病毒发病机制中经常被牵涉。卡波西肉瘤相关疱疹病毒（KSHV）与几种人类恶性肿瘤有关。它可以在潜伏感染期间稳定 HIF-1α，并在缺氧应激下经历裂解复制。然而，KSHV 通过调节 HIF-1α来控制其潜伏和裂解生命周期的机制尚未完全理解。我们之前的研究表明，缺氧敏感的染色质重塑剂 KAP1 被 KSHV 编码的潜伏相关核抗原（LANA）靶向，以抑制主要裂解复制和转录激活剂（RTA）的表达。在这里，我们进一步报告，在 KSHV 感染的原发性渗出性淋巴瘤（PEL）细胞中，基于 RNA 干扰的 KAP1 敲低会破坏病毒外显子稳定性，并取消细胞周期的 sub-G1/G1 阻滞，同时通过缺氧或使用化学 12-O-十四烷酰佛波醇-13-乙酸酯（TPA）或丁酸钠（NaB）提高 KSHV 裂解再激活的效率。此外，KSHV 全基因组筛选显示，四个缺氧反应簇在同一基因启动子内具有高 RBP-Jκ 和 HIF-1α结合位点（RBS+HRE）的一致性，并且与 KAP1 紧密相关。KAP1 的抑制极大地增强了 RBP-Jκ 与 HIF-1α 复合物的结合，从而不仅在常氧条件下，而且在缺氧条件下驱动 RTA 的表达。这些结果表明，KAP1 和 RTA 启动子内 RBS+HRE 的并存对于 KSHV 潜伏和缺氧诱导的裂解再激活都是必不可少的。
卡波西肉瘤相关疱疹病毒（KSHV）是一种 DNA 肿瘤病毒，是与几种人类恶性肿瘤相关的病原体，包括卡波西肉瘤（KS）和原发性渗出性淋巴瘤（PEL）。缺氧诱导因子 1α（HIF-1α）是一种关键的缺氧诱导因子，在 KSHV 潜伏感染的肿瘤细胞中经常升高，并有助于 KSHV 在缺氧条件下的裂解复制。然而，KSHV 通过调节 HIF-1α 来控制其潜伏和裂解生命周期的分子机制仍不清楚。在这项研究中，我们发现，在 KSHV 感染的 PEL 细胞中抑制缺氧敏感的染色质重塑剂 KAP1 会导致病毒基因组丢失，并增加其对缺氧应激的敏感性，从而导致 KSHV 裂解再激活。重要的是，我们还发现，KSHV 基因组内的四个缺氧反应簇含有高 RBP-Jκ（一种参与 Notch 信号的关键细胞调节剂）和 HIF-1α 结合位点的高一致性。这些位点也与 KAP1 紧密相关。这一发现表明，KAP1、RBP-Jκ 和 HIF-1α 通过依赖于感染细胞中氧水平的微妙相互作用，在 KSHV 发病机制中发挥着重要作用。
