RNA-mediated mechanisms of regulation of the Androgen Receptor activity in response to androgen deprivation

Androgen deprivation therapy (ADT), also called chemical castration, is one of the prevailing treatments of local and metastatic prostate cancer enabling inhibition of the Androgen Receptor (AR) pathway and tumor regression. However, the response to therapy varies among patients with some showing symptomatic relief and prolonged survival, while others relapsing in less than 5 years or failing to respond (reviewed in Harris et al, 2009). These clinical trajectories are in part due to emergence of mechanisms allowing maintenance of the AR activity, often observed in CRPC tumors, and progressive cell reprograming enabling androgen-independent growth. Activation and repression of gene exprssion by AR require interactions with numerous proteins, and very recently, with RNAs. Notably, Schmidt et al. identified a specific pyrimidine-rich motif within SRA1 and another SRA-like long noncoding RNA, named SLNCR, both binding directly the AR N-terminal domain in melanoma cells (Schmidt et al, 2016); (Schmidt et al, 2020). In addition to SLNCR, other RNAs, belonging to the long noncoding (lnc)RNA gene family, contain the aforementioned AR-interacting consensus motif, including a paragon lncRNA - HOTAIR that protects AR from proteolysis in the context of castration-resistant prostate cancer (Zhang et al, 2015) but also tethers AR to the GLI2 promoter to activate its transcription in renal cell carcinoma (Bai et al, 2021). The AR NTD is not the only domain reported to bind RNA. The growth arrest specific-5 gene GAS5, another lncRNA with well-known tumor suppressor functions, interacts with the LBD of AR inhibiting its transactivation activity and proliferation of prostate cancer cells (Lv et al, 2021). In this project we aimed to identify RNAs associated with AR in the androgen-dependent and independent cell using the technique of RNA immunoprecipitation of the transcription factor (RIP). Overall design: prostate cancer adenocarcinoma cells LNCaP growing in the presence and absence of androgen were treated with low doses of formaldehyde and 10^7 cells per condition were collected for lysis and immunoprecipitation of AR (G Hendrickson et al, 2016) .Associated RNAs were recovered and analyzed by total stranded RNA-sequencing. THe experiment was performed in duplicates and AR immunoprecipitation was compared to IgG.