PVT1 signals an androgen-dependent transcriptional repression program in prostate cancer cells and a set of the repressed genes predicts high-risk tumors

Abstract

Background Androgen receptor (AR) and polycomb repressive complex 2 (PRC2) are known to co-occupy the loci of genes that are downregulated by androgen-stimulus. Long intergenic non-coding RNA (lincRNA) PVT1 is an overexpressed oncogene that is associated with AR in LNCaP prostate cancer cells, and with PRC2 in HeLa and many other types of cancer cells. The possible involvement of PVT1 in mediating androgen-induced gene expression downregulation in prostate cancer has not been explored. Methods LNCaP cell line was used. Native RNA-binding-protein immunoprecipitation with anti-AR or anti-EZH2 was followed by RT-qPCR with primers for PVT1. Knockdown of PVT1 with specific GapmeRs (or a control with scrambled GapmeR) was followed by differentially expressed genes (DEGs) determination with Agilent microarrays and with Significance Analysis of Microarrays statistical test. DEGs were tested as a tumor risk classifier with a machine learning Random Forest algorithm run with gene expression data from all TCGA-PRAD (prostate adenocarcinoma) tumors as input. ChIP-qPCR was performed for histone marks at the promoter of one DEG. Results We show that PVT1 knockdown in androgen-stimulated LNCaP cells caused statistically significant expression upregulation/downregulation of hundreds of genes. Interestingly, PVT1 knockdown caused upregulation of 160 genes that were repressed by androgen, including a significantly enriched set of tumor suppressor genes, and among them FAS, NOV/CCN3, BMF, HRK, IFIT2, AJUBA, DRAIC and TNFRSF21. A 121-gene-set (out of the 160) was able to correctly predict the classification of all 293 intermediate- and high-risk TCGA-PRAD tumors, with a mean ROC area under the curve AUC = 0.89 ± 0.04, pointing to the relevance of these genes in cancer aggressiveness. Native RIP-qPCR in LNCaP showed that PVT1 was associated with EZH2, a component of PRC2. PVT1 knockdown followed by ChIP-qPCR showed significant epigenetic remodeling at the enhancer and promoter regions of tumor suppressor gene NOV, one of the androgen-repressed genes that were upregulated upon PVT1 silencing. Conclusions Overall, we provide first evidence that PVT1 was involved in signaling a genome-wide androgen-dependent transcriptional repressive program of tumor suppressor protein-coding genes in prostate cancer cells. Identification of transcriptional inhibition of tumor suppressor genes by PVT1 highlights the pathway to the investigation of mechanisms that lie behind the oncogenic role of PVT1 in cancer.

Unravelling similarities and differences in the role of circular and linear PVT1 in cancer and human disease

“…These PVT1 isoforms are transcribed by different promoters [63], therefore, they have to be considered separate transcriptional entities although possibly interconnected. lncPVT1 is enriched in the nucleus versus the cytosol [19,65,66], as observed by subcellular fractionation and subsequent RT-qPCR [19,[65][66][67][68], and RNA fluorescence in situ hybridisation [69,70] in several cancer cell lines. Interestingly, lncPVT1 is described as a chromatin modifier [71].…”

Identification of long noncoding RNAs in injury-resilient and injury-susceptible mouse retinal ganglion cells

“…Alternatively, lncRNAs can regulate nuclear positioning of enhancer, potentiating the enhancer to induce expression of the target genes. Others have shown that distinct lncRNAs interact with the chromatin remodeler PRC2 complex, resulting in methylation of histone H3 at lysine 27 (H3K27me3) and repression of gene transcription [ 44 – 47 ].…”

The Role of Long Non-Coding RNAs in Epithelial-Mesenchymal Transition-Related Signaling Pathways in Prostate Cancer

“…PVT1 is a widely reported lncRNA in PCa which is over-expressed and related to an unwilling prognosis ( Yang et al, 2017 ; Liu et al, 2021 ). Videria et al ( Videira et al, 2021 ) found that PVT1 may participate in an AR-dependent transcriptional repression program in PCa cells. They demonstrated that 160 genes repressed by AR were up-regulated and the epigenetic markers such as H3K27me3 and H3K27ac were remodeled when PVT1 was silenced.…”