Our previous works identified nuclear endothelial NOS (eNOS) as partner of both Estrogen Receptor Beta and Hypoxia Inducible Factors in Prostate Cancer (PCa). These transcriptional complexes determined localized chromatin remodeling in response to estrogen and hypoxia stimuli that in turn regulate genes associated with adverse prognosis in 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 associated with poor outcome and from metastatic LNCaP cells, before and after treatment with 17b-estradiol (E2). By this approach, we have documented the genome-wide existence of a considerable number of eNOS-DNA associations that define transcriptional active regions modulated by estrogen. In summary, 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. In particular, we identified of novel positive feedback loop of eNOS involving transcriptional downregulation of pri-miR34a mediated by an estrogen dependent eNOS/SIRT1 complex, as consequence we observed an up-regulation of mir-34a-target SIRT1 that in turn actives eNOS itself (by a post-transduction modification). These findings reveal unprecedented functions of eNOS and of eNOS/SIRT1 interplay, fine-tuned by E2-activated ER signaling, and attributed to eNOS a critical molecular role in aggressive PCa.
Genome-wide analysis of estrogen-dependent eNOS signaling by ChIP-sequencing: novel transcriptional mechanism in aggressive prostate cancer
Aurora Aiello;Claudia Colussi;Antonella Farsetti
2013
Abstract
Our previous works identified nuclear endothelial NOS (eNOS) as partner of both Estrogen Receptor Beta and Hypoxia Inducible Factors in Prostate Cancer (PCa). These transcriptional complexes determined localized chromatin remodeling in response to estrogen and hypoxia stimuli that in turn regulate genes associated with adverse prognosis in 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 associated with poor outcome and from metastatic LNCaP cells, before and after treatment with 17b-estradiol (E2). By this approach, we have documented the genome-wide existence of a considerable number of eNOS-DNA associations that define transcriptional active regions modulated by estrogen. In summary, 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. In particular, we identified of novel positive feedback loop of eNOS involving transcriptional downregulation of pri-miR34a mediated by an estrogen dependent eNOS/SIRT1 complex, as consequence we observed an up-regulation of mir-34a-target SIRT1 that in turn actives eNOS itself (by a post-transduction modification). These findings reveal unprecedented functions of eNOS and of eNOS/SIRT1 interplay, fine-tuned by E2-activated ER signaling, and attributed to eNOS a critical molecular role in aggressive PCa.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
