Department of Experimental Oncology, National Cancer Institute Regina Elena, Rome, Italy.
PLoS One. 2013 May 3;8(5):e62522. doi: 10.1371/journal.pone.0062522. Print 2013.
In previous work we have documented the nuclear translocation of endothelial NOS (eNOS) and its participation in combinatorial complexes with Estrogen Receptor Beta (ERβ) and Hypoxia Inducible Factors (HIFs) that determine localized chromatin remodeling in response to estrogen (E2) and hypoxia stimuli, resulting in transcriptional regulation of genes associated with adverse prognosis in prostate cancer (PCa). To explore the role of nuclear eNOS in the acquisition of aggressive phenotype in PCa, we performed ChIP-Sequencing on chromatin-associated eNOS from cells from a primary tumor with poor outcome and from metastatic LNCaP cells. We found that: 1. the eNOS-bound regions (peaks) are widely distributed across the genome encompassing multiple transcription factors binding sites, including Estrogen Response Elements. 2. E2 increased the number of peaks, indicating hormone-dependent eNOS re-localization. 3. Peak distribution was similar with/without E2 with ≈ 55% of them in extragenic DNA regions and an intriguing involvement of the 5' domain of several miRs deregulated in PCa. Numerous potentially novel eNOS-targeted genes have been identified suggesting that eNOS participates in the regulation of large gene sets. The parallel finding of downregulation of a cluster of miRs, including miR-34a, in PCa cells associated with poor outcome led us to unveil a molecular link between eNOS and SIRT1, an epigenetic regulator of aging and tumorigenicity, negatively regulated by miR-34a and in turn activating eNOS. E2 potentiates miR-34a downregulation thus enhancing SIRT1 expression, depicting a novel eNOS/SIRT1 interplay fine-tuned by E2-activated ER signaling, and suggesting that eNOS may play an important role in aggressive PCa.
在之前的工作中，我们记录了内皮型一氧化氮合酶（eNOS）的核转位及其与雌激素受体β（ERβ）和缺氧诱导因子（HIFs）形成组合复合物，这些复合物决定了对雌激素（E2）和缺氧刺激的局部染色质重塑，从而导致与前列腺癌（PCa）不良预后相关基因的转录调控。为了探讨核 eNOS 在 PCa 获得侵袭表型中的作用，我们对来自预后不良的原发肿瘤细胞和转移性 LNCaP 细胞中与染色质相关的 eNOS 进行了 ChIP-Seq 分析。我们发现：1. eNOS 结合区域（峰）广泛分布于基因组中，包含多个转录因子结合位点，包括雌激素反应元件。2. E2 增加了峰的数量，表明 eNOS 激素依赖性重新定位。3. 有无 E2 时峰的分布相似，约 55%位于外显子 DNA 区域，几个在 PCa 中失调的 miR 的 5' 结构域存在有趣的参与。已经鉴定出许多潜在的新型 eNOS 靶向基因，表明 eNOS 参与了大量基因的调控。在与不良预后相关的 PCa 细胞中，miR-34a 等一组 miR 的下调与平行发现相关，导致我们揭示了 eNOS 与 SIRT1 之间的分子联系，SIRT1 是衰老和肿瘤发生的表观遗传调节剂，受 miR-34a 负调控，反过来又激活 eNOS。E2 增强了 miR-34a 的下调，从而增强了 SIRT1 的表达，描绘了一种由 E2 激活的 ER 信号转导精细调节的新型 eNOS/SIRT1 相互作用，并表明 eNOS 可能在侵袭性 PCa 中发挥重要作用。
