Dynamic phase separation of the androgen receptor and its coactivators to regulate gene expression

Abstract

Numerous cancers, including prostate cancer (PCa), are addicted to transcription programs driven by superenhancers (SEs). The transcription of genes at SEs is enabled by the formation of phase-separated condensates by transcription factors and co-activators with intrinsically disordered regions. The androgen receptor (AR), main oncogenic driver in PCa, contains large disordered regions and is co-recruited with the co-activator MED1 to SEs to promote oncogenic programs. In this work, we show that dynamic AR-rich, liquid-like foci form in PCa models upon androgen stimulation and correlate with AR transcriptional activity. The co-activator MED1 plays an essential role in the formation of AR foci while AR antagonists hinder their formation. These results suggest that enhanced compartmentalization of AR and co-activators at SEs may play an important role in the activation of oncogenic transcription programs in PCa. A better understanding of the assembly and the regulation of these AR-rich compartments may provide novel therapeutic options.

DNA‐binding domain as the minimal region driving RNA‐dependent liquid–liquid phase separation of androgen receptor

“…As full‐length AR appears to be required for LLPS, 14 we tested its different regions for LLPS in the presence of 0.05 μg/μl poly U RNA mimic by following turbidity of the solution (optical density 600 nm OD600). The C‐terminal region of NTD (F5) predicted as a potential LLPS‐driving region (Figure 1(a), (b)), did not show an increase in turbidity, however, DBD and a longer construct F5‐DBD exhibited a transient increase in OD600 (Figure 3(a), Figure S2).…”

“…An intriguing question about the RNA/DBD biomolecular condensate formation was centered around the colocalization of DNA and RNA, as the original observation of AR LLPS demonstrated chromatin localization, 14 primarily compatible with LLPS centered around interaction with DNA. As we observed RNA‐driven phase separation, we next examined whether DNA and RNA can compete for the same binding site, and maybe form exclusive condensates (i.e., separate DNA‐DBD and RNA‐DBD droplets).…”

“…Recently, a new angle in AR research has taken shape, by the recognition that AR forms transcriptionally active foci in cells, possibly by a mechanism of LLPS in a Med1‐ 14 or SPOP‐ 15 dependent manner. LLPS of AR fits into the general trend of the formation of super enhancers by transcription factors around transcriptionally active sites in the chromatin, and may provide novel mechanistic insight leading to successful targeting of AR in PC.…”

“…MED1, a subunit of the Mediator complex that contains IDRs can phase separate on its own as well as with many TFs, including GR and ER. Recently, it was shown that androgen receptor (AR) forms dynamic AR‐rich, liquid‐like foci in cellular prostate cancer (PC) models in a MED1‐dependent manner 14 . AR phase separation was also observed to be promoted by a SPOP, a cullin3‐RING ubiquitin ligase (CRL3) adaptor of tumor suppressor activity 15 …”

“…Conventionally, drugs have been used to target the LBD, however, these are often ineffective in late—castration resistant—stages of PC, primarily because of the appearance of splice variants such as AR‐v7 that lack the ligand‐binding domain 17 . Interestingly, AR‐v7 does not undergo foci formation, 14 suggesting an internal interlay between AR domains inhibitory to LLPS.…”