Fels Institute for Cancer Research and Molecular Biology, Lewis Katz School of Medicine at Temple University, Philadelphia, Pennsylvania, United States of America.
Cancer Biology Program, Fox Chase Cancer Center, Philadelphia, Pennsylvania, United States of America.
PLoS Pathog. 2018 Nov 5;14(11):e1007394. doi: 10.1371/journal.ppat.1007394. eCollection 2018 Nov.
Latent membrane protein 1 (LMP1) is the major transforming protein of Epstein-Barr virus (EBV) and is critical for EBV-induced B-cell transformation in vitro. Poly(ADP-ribose) polymerase 1 (PARP1) regulates accessibility of chromatin, alters functions of transcriptional activators and repressors, and has been directly implicated in transcriptional activation. Previously we showed that LMP1 activates PARP1 and increases Poly(ADP-ribos)ylation (PARylation) through PARP1. Therefore, to identify targets of LMP1 that are regulated through PARP1, LMP1 was ectopically expressed in an EBV-negative Burkitt's lymphoma cell line. These LMP1-expressing cells were then treated with the PARP inhibitor olaparib and prepared for RNA sequencing. The LMP1/PARP targets identified through this RNA-seq experiment are largely involved in metabolism and signaling. Interestingly, Ingenuity Pathway Analysis of RNA-seq data suggests that hypoxia-inducible factor 1-alpha (HIF-1α) is an LMP1 target mediated through PARP1. PARP1 is acting as a coactivator of HIF-1α-dependent gene expression in B cells, and this co-activation is enhanced by LMP1-mediated activation of PARP1. HIF-1α forms a PARylated complex with PARP1 and both HIF-1α and PARP1 are present at promoter regions of HIF-1α downstream targets, leading to accumulation of positive histone marks at these regions. Complex formation, PARylation and binding of PARP1 and HIF-1α at promoter regions of HIF-1α downstream targets can all be attenuated by PARP1 inhibition, subsequently leading to a buildup of repressive histone marks and loss of positive histone marks. In addition, LMP1 switches cells to a glycolytic 'Warburg' metabolism, preferentially using aerobic glycolysis over mitochondrial respiration. Finally, LMP1+ cells are more sensitive to PARP1 inhibition and, therefore, targeting PARP1 activity may be an effective treatment for LMP1+ EBV-associated malignancies.
潜伏膜蛋白 1（LMP1）是 Epstein-Barr 病毒（EBV）的主要转化蛋白，对于 EBV 诱导的体外 B 细胞转化至关重要。聚（ADP-核糖）聚合酶 1（PARP1）调节染色质的可及性，改变转录激活因子和转录抑制因子的功能，并且直接参与转录激活。之前我们表明，LMP1 通过 PARP1 激活 PARP1 并增加聚（ADP-核糖）化（PARylation）。因此，为了鉴定通过 PARP1 调节的 LMP1 靶标，我们在 EBV 阴性 Burkitt 淋巴瘤细胞系中外源性表达 LMP1。然后用 PARP 抑制剂奥拉帕利处理这些表达 LMP1 的细胞，并进行 RNA 测序。通过这项 RNA-seq 实验鉴定的 LMP1/PARP 靶标主要涉及代谢和信号转导。有趣的是，RNA-seq 数据的 Ingenuity 通路分析表明，缺氧诱导因子 1-α（HIF-1α）是通过 PARP1 介导的 LMP1 靶标。PARP1 作为 B 细胞中 HIF-1α 依赖性基因表达的共激活因子发挥作用，并且这种共激活作用通过 LMP1 介导的 PARP1 激活得到增强。HIF-1α 与 PARP1 形成 PARylated 复合物，HIF-1α 和 PARP1 均存在于 HIF-1α 下游靶标启动子区域，导致这些区域的正组蛋白标记物积累。PARP1 抑制可减弱复合物形成、PARylation 以及 PARP1 和 HIF-1α 在 HIF-1α 下游靶标启动子区域的结合，随后导致抑制性组蛋白标记物的积累和正组蛋白标记物的丢失。此外，LMP1 将细胞切换为糖酵解的“Warburg”代谢，优先使用有氧糖酵解而不是线粒体呼吸。最后，LMP1+细胞对 PARP1 抑制更敏感，因此，靶向 PARP1 活性可能是治疗 LMP1+ EBV 相关恶性肿瘤的有效方法。
