Global analysis of transcription in castration-resistant prostate cancer cells uncovers active enhancers and direct androgen receptor targets

Abstract

Androgen receptor (AR) is a male sex steroid-activated transcription factor (TF) that plays a critical role in prostate cancers, including castration-resistant prostate cancers (CRPC) that typically express amplified levels of the AR. CRPC-derived VCaP cells display an excessive number of chromatin AR-binding sites (ARBs) most of which localize to distal inter- or intragenic regions. Here, we analyzed direct transcription programs of the AR in VCaP cells using global nuclear run-on sequencing (GRO-seq) and integrated the GRO-seq data with the ARB and VCaP cell-specific TF-binding data. Androgen immediately activated transcription of hundreds of protein-coding genes, including IGF-1 receptor and EGF receptor. Androgen also simultaneously repressed transcription of a large number of genes, including MYC. As functional enhancers have been postulated to produce enhancer-templated non-coding RNAs (eRNAs), we also analyzed the eRNAs, which revealed that only a fraction of the ARBs reside at functional enhancers. Activation of these enhancers was most pronounced at the sites that also bound PIAS1, ERG and HDAC3, whereas binding of HDAC3 and PIAS1 decreased at androgen-repressed enhancers. In summary, our genome-wide data of androgen-regulated enhancers and primary target genes provide new insights how the AR can directly regulate cellular growth and control signaling pathways in CPRC cells.

Androgen receptor enhancer usage and the chromatin regulatory landscape in human prostate cancers

“…A further elucidation of direct AR target genes has been performed by combining AR chromatin-binding profiles with gene expression profiling. Upon androgen stimulation, hundreds to thousands of genes are being either up-or downregulated, dependent on hormone stimulation times and bioinformatics cut-offs used (DePrimo et al 2002, Hendriksen et al 2006, Massie et al 2011, Toropainen et al 2016. Apart from protein-coding genes, non-coding transcripts are also under direct control of AR (Wang et al 2011, Toropainen et al 2016.…”

“…Of note, the number of genome-wide AR-binding sites is considerably higher than the number of differentially expressed genes upon androgen stimulation. Moreover, merely 30% of the AR-bound enhancers show eRNA production (Toropainen et al 2016). This suggests that only a fraction of AR-bound enhancers are functional or that these enhancers can function without eRNA transcription.…”

Nascent transcript analysis of glucocorticoid crosstalk with TNF defines primary and cooperative inflammatory repression

“…It has been argued that redistribution of limiting amounts of specific co-regulators from one set of enhancers to another is a unifying explanation for rapid repression in association with transcriptional induction (Guertin et al 2014;Schmidt et al 2016), which has been observed in a variety of other signaling contexts (Hah et al 2011;Step et al 2014;Franco et al 2015;Loft et al 2015;Toropainen et al 2016). Although redistribution of FOXA1 occupancy with glucocorticoid treatment has been reported (Swinstead et al 2016), whether specific transcription factors or co-activators rapidly redistribute their nuclear position from repressed to activated regulatory elements in association with GR activation has not been fully investigated.…”

Interplay between the androgen receptor signaling axis and microRNAs in prostate cancer

“…However, it is often difficult to identify which of these transcripts are directly regulated by AR, largely due to the fact that AR often binds to enhancers that are distal to its target genes (Wang et al 2007). Carefully assessing the temporality of androgenmediated gene regulation (Massie et al 2011), integrating transcriptomic data with AR cistromes (Pomerantz et al 2015) and/or using more sophisticated transcriptomic techniques such as global run-on sequencing (GRO-seq) (Toropainen et al 2016), which is designed to identify nascent transcription, have improved the detection of bona fide AR targets in PCa cell lines and tissues. Nevertheless, accurately differentiating direct vs indirect targets of the AR remains challenging.…”